mutágenos en suelos contaminados
TRANSCRIPT
www.elsevier.com/locate/reviewsmr
Mutation Research 567 (2004) 227–345
ReviewCommunity address: www.elsevier.com/locate/mutres
Mutagens in contaminated soil: a review
Paul A. Whitea,*, Larry D. Claxtonb
aMutagenesis Section, Safe Environments Program, Health Canada, Tunney’s Pasture 0803A, Ottawa, Ont., Canada K1A 0L2bEnvironmental Carcinogenesis Division (MD-B143-06), US Environmental Protection Agency,
Research Triangle Park, NC 27711, USA
Available online 18 November 2004
Abstract
The intentional and accidental discharges of toxic pollutants into the lithosphere results in soil contamination. In some cases
(e.g., wood preserving wastes, coal-tar, airborne combustion by-products), the contaminated soil constitutes a genotoxic hazard.
This work is a comprehensive review of published information on soil mutagenicity. In total, 1312 assessments of genotoxic
activity from 118 works were examined. The majority of the assessments (37.6%) employed the Salmonella mutagenicity test
with strains TA98 and/or TA100. An additional 37.6% of the assessments employed a variety of plant species (e.g., Tradescantia
clone 4430, Vicia faba, Zea mays, Allium cepa) to assess mutagenic activity. The compiled data on Salmonella mutagenicity
indicates significant differences (p < 0.0001) in mean potency (revertents per gram dry weight) between industrial, urban, and
rural/agricultural sites. Additional analyses showed significant empirical relationships between S9-activated TA98 mutagenicity
and soil polycyclic aromatic hydrocarbon (PAH) concentration (r2 = 0.19 to 0.25, p < 0.0001), and between direct-acting TA98
mutagenicity and soil dinitropyrene (DNP) concentration (r2 = 0.87, p < 0.0001). The plant assay data revealed excellent
response ranges and significant differences between heavily contaminated, industrial, rural/agricultural, and reference sites, for
the anaphase aberration in Allium cepa (direct soil contact) and the waxy locus mutation assay in Zea mays (direct soil contact).
The Tradescantia assays appeared to be less responsive, particularly for exposures to aqueous soil leachates. Additional data
Abbreviations: Ac, acetone; ACN, acetonitrile; BaP, benzo(a)pyrene; Bq, Becquerel; CA, chromosomal aberration; ASTM, American
Society for Testing and Materials; CERCLA, Comprehensive Environmental Response, Compensation, and Liability Act; CERCLIS,
Comprehensive Environmental Response, Compensation, and Liability Information System; Cyclohex, cyclohexane; DCM, dichloromethane;
DDE, 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene; DEK, diethylketone; DMSO, dimethylsulfoxide; DNP, dinitropyrene; EDC, ethylene
dichloride; EPCRA, Emergency Planning and Community Right-To-Know Act; EtOH, ethanol; GAP, Genetic Activity Profile; HDPE, high
density polyethylene; Hex, hexane; HMX, cyclotetramethylene-tetranitramine; HPRT, hypoxanthine phosphoribosyltransferase; IARC, Inter-
national Agency for Research on Cancer; MetOH, methanol; MN, micronucleus; NFRAP, No Further Remedial Action Planned; NPL, National
Priorities List; NPRI, National Pollutant Release Inventory; PAH, polycyclic aromatic hydrocarbon; PCB, polychlorinated biphenyl; PCDD,
polychlorinated dibenzodioxin; PCDF, polychlorinated dibenzofuran; PCP, pentachlorophenol; Pfu, plaque forming unit; PPA, Pollution
Prevention Act; Prop, 2-propanol; RCRA, Resource Conservation and Recovery Act; RDX, hexahydro-1,3,5-trinitro-1,3,5-triazine; Rev,
Salmonella revertent; SCE, sister chromatid exchange; SDR, studentized deleted residual; SHM, stamen hair mutation; SIC, Standard Industrial
Classification; S.E., standard error of the mean; Tol, toluene; TNT, trinitrotoluene; Trad, Tradescantia; TRI, Toxic Release Inventory; WPW,
wood preserving waste
* Corresponding author. Tel.: +1 613 941 7373; fax: +1 613 941 8530.
E-mail address: [email protected] (P.A. White).
1383-5742/$ – see front matter. Crown Copyright # 2004 Published by Elsevier B.V. All rights reserved.
doi:10.1016/j.mrrev.2004.09.003
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345228
analyses showed empirical relationships between anaphase aberrations in Allium, or mutations in Arabidopsis, and the 137Cs
contamination of soils. Induction of micronuclei in Tradescantia is significantly related to the soil concentration of several
metals (e.g., Sb, Cu, Cr, As, Pb, Cd, Ni, Zn). Review of published remediation exercises showed effective removal of genotoxic
petrochemical wastes within one year. Remediation of more refractory genotoxic material (e.g., explosives, creosote) frequently
showed increases in mutagenic hazard that remained for extended periods. Despite substantial contamination and mutagenic
hazards, the risk of adverse effect (e.g., mutation, cancer) in humans or terrestrial biota is difficult to quantify.
Crown Copyright # 2004 Published by Elsevier B.V. All rights reserved.
Keywords: Salmonella; Tradescantia; Allium; Mutagen; Clastogen; Soil pollution
Contents
1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229
2. Sources of soil contamination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229
3. Investigating genotoxic substances in soil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 233
3.1. The nature of soil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 233
3.2. Detection of mutagens in soil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 237
3.3. Bioassays employed for soil mutagenicity assessment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 238
4. Collection and analysis of published soil mutagenicity/clastogenicity data . . . . . . . . . . . . . . . . . . . . . . . . . . 239
5. Salmonella mutagenicity of soils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 241
5.1. TA98 and TA100 mutagenicity of soil extracts. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 242
5.2. Relationships between Salmonella mutagenicity and extraction solvent . . . . . . . . . . . . . . . . . . . . . . . . 247
5.3. Relationships between Salmonella mutagenicity and soil contamination . . . . . . . . . . . . . . . . . . . . . . . 248
5.4. Mutagenic activity detected using other Salmonella strains . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 250
5.5. Miscellaneous Salmonella mutagenicity data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 252
6. Other prokaryote or molecular in vitro assays used for soil genotoxicity assessment . . . . . . . . . . . . . . . . . . . 252
7. Plant assays used for soil genotoxicity assessment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 258
7.1. Allium cepa anaphase aberration data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 259
7.2. Arabidopsis gene mutation data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 261
7.3. Zea mays waxy locus mutation data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 263
7.4. Tradescantia stamen hair mutation data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 265
7.5. Tradescantia micronucleus test data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 267
7.6. Other plant assays used for soil mutagenicity assessment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 270
8. Other eukaryotic assays used for soil genotoxicity assessment. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 273
8.1. In vitro eukaryotic assays . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 273
8.2. In vivo eukaryote assays . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 277
9. Miscellaneous soil genotoxicity results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 279
10. Remediation of genotoxic soils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 279
11. Summary and discussion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 285
12. Conclusions and future prospects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 293
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 229
Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 294
Appendix A: Salmonella mutagenicity data collected from the literature . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 295
Appendix B: Plant clastogenicity and mutagenicity data collected from the literature . . . . . . . . . . . . . . . . . . . . . 315
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 330
1. Introduction
An unfortunate consequence of industrialization and
industrial production is the generation and release of
toxic waste products. Although these wastes can be
treated, reused and recycled, large quantities of toxic
material is released into the atmosphere, hydrosphere
and lithosphere. Although much of the waste is released
directly into the atmosphere and hydrosphere (i.e.,
surface waters of lakes, rivers, and streams), land
disposal activities, intentional or otherwise, contribute
to direct contamination of surface soils and subterra-
nean (groundwater) aquifers. Intentional land disposal
activities such as landfills, lagoons, surface impound-
ments, ponds, septic systems, and land treatment are
cost effective disposal strategies that take advantage of
the enormous capacity of soil to retain and degrade
toxic pollutants. However, inadequate information
about waste toxicity and post-disposal behaviour, poor
planning, improper disposal, and poor management of
disposal sites has resulted in serious contamination
problems at industrial and hazardous waste disposal
sites. Moreover, accidental leaks/spills occurring
during transport and storage of industrial materials
(e.g., solvents, fuels, etc.) have resulted in contamina-
tion problems at sites not intended for waste disposal.
Finally, widespread fossil fuel combustion, solid waste
incineration, and pesticide application has also con-
tributed to regional soil contamination.
Prior to the mid-1970s few countries had regulations
restricting land disposal of hazardous waste materials.
Consequently, it was difficult to estimate the numbers of
contaminated industrial areas and hazardous waste
disposal sites, the total area of contaminated land in a
given country, or the approximate liability and
remediation costs associated with these sites. However,
a gradual increase in environmental awareness, and
evidence of serious mismanagement of industrial and
waste disposal sites, has prompted many countries to
pass legislation aimed at proper management and
rehabilitation of present and future hazardous waste
disposal areas. For example, in 1976 the United States
passed the Resource Conservation and Recovery Act
(RCRA), followed in 1980 by the Comprehensive
Environmental Response, Compensation, and Liability
Act (CERCLA), otherwise known as the Superfund
Act. These acts ensured proper management and
permitting of waste disposal sites. Moreover, they
require an inventory and assessment of older sites
including orphaned sites resulting from facility closure
and site abandonment. As of May 2004 the US
Comprehensive Environmental Response, Compensa-
tion, and Liability Information System (CERCLIS)
contained information on 45,516 contaminated sites,
including 12,040 Superfund sites currently under
investigation and 33,476 NFRAP (no further remedial
action planned) sites that have been removed from the
Superfund list. The Superfund list includes 1238
National Priorities List (NPL) sites that have been
thoroughly evaluated and warrant further investigation
to assess exposure pathways and the extent of human
health or environmental hazards. This information, in
addition to an inventory of contaminated sites in
Canada and several European nations, is provided in
Table 1. The data presented indicate that the United
States and Canada collectively possess more than
60,000 contaminated sites. Moreover, several indus-
trialized countries in Western Europe that are only a
fraction of the size of the USA or Canada (e.g.,
Germany, Denmark, Switzerland, Austria) have equiva-
lent or greater numbers of contaminated sites.
2. Sources of soil contamination
The quantities of toxic industrial wastes released to
land vary according to industrial sector. Until recently,
the exact annual land releases of toxic material were
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345230
Table 1
Inventory of contaminated terrestrial sites in several industrialized countries
Country Total
area (km2)
Population
(millions)
Number of
contaminated sitesa
Sites per
1000 km2
Sites per 106
people
Canada 9,976,140 31.6 15,000–40,000 1.5–4.0 475–1266Federal sites 3,843 0.4 121.6
United States 9,656,345 288.4 45,516 4.7 157.8Total CERCLISb sites 12,040 1.2 41.8
Federal CERCLIS sites 1,021 0.1 3.5
Total NFRAPc sites 33,476 3.5 116.1
Federal NFRAP sites 1,032 0.1 3.6
Current NPLd sites 1,238 0.1 4.3
Germanye 356,910 82.8 202,880 568.4 2450.2Denmark 43,090 5.3 37,000 858.7 6981.1Switzertland 41,290 7.3 35,000 847.7 4794.5Austria 83,850 8.1 28,000 333.9 3456.8Finland 338,130 5.2 10,396 30.7 1999.2Italy 301,270 57.6 8,873 29.5 154.0Belgium 30,518 10.2 7,728 253.2 757.6Sweden 449,960 8.9 7,000 15.6 786.5Spain 504,780 40.0 4,902 9.7 122.6Norway 323,900 4.5 2,121 6.5 471.3Lithuania 65,300 3.6 4,430 67.8 1230.6Romania 238,381 22.4 1,634 6.9 72.9Estonia 45,227 1.4 1,565 34.6 1117.9
a Values in bold are totals for each country. Values for Canada are estimates from the Ministry of Environment or from The Federal
Contaminated Sites and Solid Waste Landfills Inventory (http://publiservice.tbs-sct.gc.ca/dfrp-rbif/cs-sc/home-accueil.asp?Language=EN).
Values for the USA refer to identified contaminated sites as of May 2004 (see http://www.epa.gov/superfund/sites/phonefax/products.htm).
Values for western European countries refer to identified sites of potential contamination [452]. Values for former Warsaw Pact countries refer to
contaminated sites that have been registered with the relevant authorities [453].b The Comprehensive Environmental Response, Compensation, and Liability Act (CERCLA) Information System, a database of current
Superfund sites.c Archived ‘‘No Further Remedial Action Planned’’ sites that have been removed from the Superfund list.d National Priorities List. Prioritized Superfund sites slated for hazard ranking and remedial action.e Potentially contaminated current and former military sites not included.
difficult, if not impossible, to determine. However, in
response to catastrophes such as the industrial accident
in Bhopal, India in 1984 many countries began tracking
the production, usage, and release of toxic substances.
For example, in 1986 the US government passed the
Emergency Planning and Community Right-To-Know
Act (EPCRA). This legislation was enacted to promote
emergency planning, and provide the public with
information on the use, production, and release of toxic
substances. To accomplish this monumental informa-
tion distribution task the EPCRA, and the subsequent
Pollution Prevention Act (PPA), required the creation of
a publicly accessible database containing information
on the release of toxic chemicals. This database is
known as the Toxics Release Inventory (TRI). Several
countries followed with similar databases, such as
Canada’s National Pollutant Release Inventory (NPRI)
established in the early 1990s as part of the Canadian
government’s Green Plan.
The 2002 TRI data indicate that a total of 24,379
industrial facilities released 2.17 billion kg of toxic
material (518 substances or substances groups).
Approximately 53%, or 1.15 billion kg of material,
was disposed via on- and off-site land release.
Therefore, in 2002 American industries disposed of
1.15 billion kg of toxic waste via on- and off-site
landfill, land treatment, and surface impoundment. The
2002 Canadian NPRI data indicate that 4596 industries
released a total of 402.5 million kg of toxic material
(274 substances). Approximately 8.1% or 32.5 mil-
lion kg was disposed via on-site land release. Therefore,
in 2002 Canadian industries disposed of 32.5 million kg
of toxic waste via on-site landfill, land treatment, and
surface impoundment. The difference between the
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 231
proportions of American and Canadian total releases
disposed via land release is primarily due to differences
in the industrial sectors monitored, and inclusion of off-
site landfill and land farming activities in the US
inventory. For example, the US release inventory
includes all metal mining facilities, whereas only
mining facilities involved in the secondary processing
of mined material are included in the Canadian
inventory. A summary of the 2002 land releases in
the US and Canada by industrial sector is provided in
Tables 2a and 2b. The data in Table 2a indicate that in
the United States over 70% of the total annual on- and
off-site land releases of 1149.1 million kg are accounted
for by metal mining and processing facilities, with
another 14% accounted for by electric, gas, and sanitary
Table 2a
2002 TRI on- and off-site land releases in the United States (by industria
Industry type
Metal mining (e.g., Fe, Cu, Pb, Zn, Au, Ag)
Primary metal smelting and processing
Electric, gas, and sanitary services
Solvent recovery operations (under RCRAc)
Chemical and allied products
Paper and allied products
Food and related products
Coal mining and coal mine services
Fabricated metal products
Transportation equipment manufacture
Plastic and rubber products
Stone, clay, glass, and concrete products
Electronic and other electrical equipment
Industrial and commercial machinery
Petroleum refining and related industries
Lumber and wood products
Leather and leather products
Textile mill products
Tobacco manufacture
Photographic, medical, and optical goods
Petroleum bulk stations and terminals
Chemical wholesalers
Apparel manufacture
Furniture and fixtures
Printing, publishing, and related industries
No reported SIC code
Miscellaneous manufacturing
Industries with multiple SIC codes
Total
a Standard Industrial Classification Codes (see http://www.osha.gov/plb Includes on- and off-site disposal via landfill, land treatment, surface im
Inventory. Available online at http://www.epa.gov/triexplorer. Values rounc The US Resource Conservation and Recovery Act.
services, primary metal processing and smelting, and
solvent recovery operations. In Canada (Table 2b),
chemical and allied industries account for over 32% of
the total annual on-site land releases of 32.5 million kg,
with a further 48% accounted for by primary metal
processing and smelting, electric, gas, and sanitary
services, and pulp and paper production.
Neither the Canadian NPRI, nor the American TRI
provides a complete list of all land releases of toxic
pollutants. For the most part, these lists contain an
inventory of releases for substances that are manu-
factured, processed, or used in fairly large quantities
(e.g., >10,000 kg for NPRI Group I substances,
�5000–10,000 kg for TRI) by fairly large industries
(>10 full-time employees). In addition, the inventories
l sector)
SICa Land releases (�106 kg)b
10 580.1
33 245.2
49 159.9
4953/7389 69.5
28 31.2
26 9.0
20 8.6
12 6.7
34 5.6
37 4.0
30 3.9
32 3.8
36 1.8
35 1.8
29 1.0
24 1.0
31 0.4
22 0.2
21 0.2
38 0.1
5171 0.1
5169 0.1
23 <0.03
25 <0.02
27 <0.01
– 5.1
39 0.3
20–39 9.6
1149.1
s/imis/sic_manual.html).
poundment, and underground injection. Source: 2002 Toxic Release
ded to the nearest 105 kg.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345232
Table 2b
2002 NPRI on-site land releases in Canada (by industrial sector)
Industry type SICa Land releases (�103 kg)b
Chemical and allied products 28 10422.6
Electric, gas, and sanitary services 49 8511.4
Primary metal smelting and processing 33 4868.2
Paper and allied products 26 2204.6
Air transportation 45 1936.4
Food and related products 20 1334.0
Heavy construction 1629 950.8
Miscellaneous services and consulting 8999 651.1
National security 9711 388.1
Crude petroleum and natural gas 1311 324.3
Lumber and wood products 24 231.1
Wholesale trade — durable goods/scrap 50 187.4
Metal mining (e.g., Fe, Cu, Pb, Zn, Au, Ag) 10 131.3
Air/water resource and solid waste Management 9511 124.6
Plastic and rubber products 30 65.5
Petroleum and coal products 29 58.6
Miscellaneous manufacturing 39 25.1
Commercial printing 2752 21.4
Stone, clay, glass, and concrete products 32 13.5
Chemical and allied products/petroleum bulk 51 3.7
Health and allied services 80 1.0
Electrical and electronic equipment 36 0.5
Correctional institutions 9223 0.4
Special trade contractors (e.g., paint removal) 1799 0.3
Crude petroleum pipelines 4612 0.1
Fabricated metal products 34 0.1
Farm machinery and equipment 3523 0.1
Aircraft parts and auxiliary equipment 3728 <0.1
Commercial research (physical and biological) 8731 <0.1
Automotive transmission repair 7537 <0.1
Total 32456.2
a Standard Industrial Codes (see http://www.osha.gov/pls/imis/sic_manual.html).b Includes on-site landfill disposal, land treatment, surface impoundment, leaks, and spills. Source: 2002 National Pollutant Release Inventory
of 266 substances. Available online at http://www.ec.gc.ca/pdb/npri/npri_dat_rep_e.cfm#complete. Criteria air pollutants (e.g., SO2, NOx, CO,
total particulates) were not included.
do not provide any indication of post-emission
behaviour or the potential human hazard of the toxic
emissions. However, both the TRI of 518 substances
and the NPRI of 274 substances do provide a separate
inventory of substances that are known or probable
animal carcinogens. Of the 518 substances monitored
by the TRI, 144 substances are described as known or
suspected animal carcinogens (i.e., IARC categories 1,
2A, and 2B). Of the 274 substances monitored by the
NPRI, only 20 known or suspected animal carcinogens,
or groups of compounds that include carcinogens (e.g.,
polycyclic aromatic hydrocarbons or PAHs), had
measurable land releases. According to The Interna-
tional Agency for Research on Cancer (IARC), these
substances are predominantly manufactured, processed
and released by 15 industrial categories: rubber
production, printing and print processing, pulp and
paper production, dry cleaning, paint manufacture and
painting, inorganic chemical production, carpentry and
joinery, leather tanning and shoe manufacture, petro-
leum refining, iron founding and coke production, coal
gasification, aluminum production, textile manufacture
and finishing, organic chemical production, and glass
manufacturing (see http://monographs.iarc.fr/ for addi-
tional details).
A survey of the 2002 TRI results indicates that total
US on- and off-site land releases of suspected or
known carcinogens was 513.8 million kg. The 2002
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 233
NPRI results indicate that total Canadian on-site land
releases of carcinogens during the same period were
3.8 million kg. In both cases, inorganic chemicals
such as asbestos, lead, chromium, nickel, and arsenic
(and related compounds) account for almost all
(�99%) of the annual carcinogenic releases. Several
of the carcinogens included in these release values,
including some substances with the highest on-site
land releases, are also well known mutagens and/or
clastogens. Examples include chromium and arsenic
compounds, asbestos, creosote, styrene, benzene,
urethane, formaldehyde, vinyl chloride, tetrachlor-
oethylene, and a variety of PAHs. Table 3a sum-
marizes the 2002 TRI on- and off-site land releases of
noteworthy mutagens, clastogens, and carcinogens
with annual releases of at least 103 kg. Table 3b
summarizes the 2002 NPRI on-site land releases of
noteworthy mutagens, clastogens, and carcinogens
with annual releases of at least 100 kg. It should be
noted that neither inventory would include atmo-
spheric fallout of mutagenic aromatic hydrocarbons
and their derivatives (e.g., PAHs, nitroarenes, quino-
lines, thiophenes, etc.) that are by-products of
industrial and domestic fossil fuel combustion, and
waste incineration [1–3]. In addition, the NPRI would
not include off-site transfers for land disposal (e.g.,
land treatment).
3. Investigating genotoxic substances in soil
3.1. The nature of soil
Soil is a dynamic and complex medium that forms
at the interface of the atmosphere, lithosphere,
hydrosphere, and biosphere. It is essentially an
aggregate of unconsolidated mineral and organic
material produced by a complex combination of
physical, chemical, and biological processes [4,5].
The properties of soil are spatially and temporally
variable, and dependent on the combined effects
climate, biological activity, topography, and the
mineralogical composition of the parent rock. On a
volumetric basis soil is about 45% mineral, 2–5%
organic material, 20–30% air, and 20–30% water
[5,6]. The bulk density of mineral soil ranges from
0.86 to 2.08 g/cm3, with typical organic soils yielding
values between 0.1 and 0.6 g/cm3 [7]. On a dry weight
basis, soils are approximately 93–99% mineral with
the remainder composed of organic material [6].
The mineral portion of soil is most often described
in terms of its particle size distribution. Soil mineral
particles can range from extremely fine clay particles
to coarse sand. The primary soil particles, which resist
further breakdown, are sand (0.5–2 mm diameter), silt
(0.002–0.5 mm diameter), and clay (<0.002 mm
diameter). The relative proportions of each of these
minerals determines the property known as soil texture
[5]. The organic material portion of soil is derived
from decomposed or partially decomposed plant and/
or animal tissues. Much of this material (85–90%)
consists of humic matter, complex high molecular
weight polymers formed from the microbial decom-
position of organic material [4]. Although most soils
contain 2–5% organic matter by volume, the values
can range from less than 1% to greater than 80%.
These highly organic soils are usually referred to as
peat.
Variations in soil texture (i.e., particle size
distribution), organic matter content, water content,
and a variety of other variables (e.g., gradation,
consistency, porosity, permeability, compressibility,
oxidation state, particle shape, particle charge, and
cation exchange capacity), results in an enormous
range of soil types with vastly different physical and
chemical properties [7]. Several soil classification
systems have been devised to categorize soils with
different properties. One of the more common textural
classification systems, devised by the US Department
of Agriculture, relies on the familiar triangular
diagram with the three apexes representing sand, silt
and clay content. Using the percent (by weight) of
clay, silt and sand, any soil sample that passes through
a 2 mm sieve can be positioned on the diagram and
classified. Detailed descriptions of the various soil
classification systems (e.g., Unified Soil Classification
System, American Association of State Highway and
Transportation Officials Classification System, US
Comprehensive Soil Classification System) that
categorize soils on the basis of texture, organic matter
content, and oxidation state are beyond the scope of
this work. For additional details on soil classification,
as well as soil chemistry and soil physics, the reader
should consult Weingardner [5] or Dragun [7].
Differences in soil type can have profound effects
on the environmental fate and toxicity of soil
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345234
Table 3a
2002 TRI on- and off-site land releases of carcinogenic substances in the United States (>1000 kg only)a
Compound Mutagenicity/clastogenicityb,c Carcinogenicityd Total land releases (�103 kg)
Arsenic compoundse +/CC 1 182033.2
Lead compoundsf CC 2B 181618.0
Chromium compoundsg +++/CC 1 19089.8
Nickel compoundsh ++/CC 1 15669.8
Asbestos (friable) C 1 3016.2
Cobalt compoundsi ++/CC 2B 2697.0
Cadmium compoundsj ++/CC 1 2396.9
Polychlorinated biphenyls � 2A 872.4
Styrene +++/CC 2B 774.4
Creosotek ++ 2A 765.1
1,4-Dioxane +l/C 2B 438.2
Polycyclic aromatic compoundsm 427.2
Benz(a)anthracene +++/CC 2A
Benzo(a)pyrene +++/CC 2A
Benzo(b)fluoranthene +/C 2B
Dibenzo(a,h)anthracene ++/CC 2A
Indeno(1,2,3-cd)pyrene + 2B
Dibenz(a,h)acridine + 2B
Beryllium compoundsn ++/C 1 362.3
Bis(2-ethylhexyl)phthalate +o/C 3 335.1
Formaldehyde +++/CC 2A 208.0
Chlorothalonil +p/C 2B 115.4
Polychlorinated alkanes +p/C 2Bq 91.2
Tetrachloroethylene +/C 2B 86.2
Dichloromethane ++/CC 2B 65.2
Vinyl acetate CC 2B 49.3
Benzene +++/CC 1 48.6
Propylene oxide +++/CC 2B 42.0
Chloroform +/CC 2B 40.6
Ethyl acrylate ++/CC 2B 39.8
Urethane (ethyl carbamate) +++/CC 2B 20.0
Ethylbenzene +p 2B 18.6
Toluene diioscyanater ++s/C 2B 15.4
Catechol ++/C 2B 14.2
1,2-Dichloroethane +++/C 2B 13.8
Trichloroethylene ++/C 2A 12.6
2,4-Dichloro-1-(4-nitrophenoxy)-benzene + 2Bt 11.5
Diaminotoluene � 2Bu 8.9
Acetaldehyde +++/CC 2B 8.4
Acrylonitrile +++/CC 2B 7.6
Hexachlorobenzene + 2B 6.9
Nitrilotriacetic acid +o 2B 6.4
4,40-Diaminodiphenyl ether + 2B 6.4
Pentachlorophenol +/CC 2B 5.1
Nitrobenzene � 2B 5.1
2,4-Dichlorophenoxyacetic acid +++/C 2Bv 3.9
Methoxone +++/CCY 2Bv 3.8
Toluene-2,4-diisocyanate ++s/C 2B 3.5
Ethylene oxide +++/CC 1 3.4
Carbon tetrachloride +/C 2B 3.0
Epichlorohydrin ++/CC 2A 2.8
4,40-Methylenedianiline ++/CC 2B 2.7
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 235
Table 3a (Continued )
Compound Mutagenicity/clastogenicityb,c Carcinogenicityd Total land releases (�103 kg)
Toxaphene +/C 2B 2.6
1,3-Butadiene +++/CC 2A 1.6
Mecoprop � 2Bv 1.5
Acrylamide +++/CC 2A 1.4
Chlorophenols + 2B 1.2
2,4-Dinitrotoluene +++ 2B 1.1
Ethylene thiourea + 2B 1.0
Total inorganic 406883.1
Total organic 4593.2
Grand total 411476.3
a Includes on- and off-site disposal via landfill, land treatment, surface impoundment, and underground injection. See Table 2a for additional
details. Thirty-one other organic compounds with values >100 and <1000 kg (i.e., chlordane, tolunene-2,6-diisocyanate, 1,2,3-trichloropro-
pane, 1,4-dichlorobenzene, 2,6-dinitrotoluene, hexachloroethane, 1,3-dichloropropylene, N-nitrosodi-N-butylamine, N-nitrosodiethylamine,
chlorendic acid, 2-nitropropane, heptachlor, vinyl chloride, thiourea, o-toluidine, dimethyl sulfate, dichlorovos, 2,4,6-trichlorophenol, 1,2-
dibromo-3-chloropropane, 1,2-dibromoethane, 2-acetlyaminofluorene, 4,40-methylene-bis(2-chloroaniline), 4-dimethylaminoazobenzene,
hydrazine, N-nitrosodi-N-proplyamine, N-nitrosopiperidine, p-chloroaniline, propane sultone, safrole, and 2,4-dichlorophenoxyacetic acid
2-ethylhexyl ester) account for an additional 7160.4 kg.b Based on data from the Genetic Activity Profile (GAP) database [454,455], the Monograph Series of the International Agency for Research
on Cancer (IARC) (available online at http://monographs.iarc.fr/), and the published results of the US national Toxicology Program (available
online at http://ntp-server.niehs.nih.gov/).c (�) Compounds for which there is no evidence of mutagenicity or clastogenicity, (+) mutagenic in bacterial and/or fungal/yeast cells in
vitro, (++) also mutagenic in plants or animal cells in vitro, (+++) also mutagenic in the Drosophila melanogaster somatic mutation and
recombination test, and/or sex-linked recessive lethal test, and/or transgenic rodent assays, and/or rodent dominant lethal test. For cytogenetic
endpoints C refers to substances that are clastogenic in in vitro assays, CC refers to substances that are clastogenic both in vitro and in vivo. Note:
In some instances conflicting results have been reported in the literature.d IARC classification system: (1) carcinogenic to humans, (2A) probably carcinogenic to humans, (2B) possibly carcinogenic to humans, (3)
inadequate or limited evidence of carcinogenicity in experimental animals.e Both the +3 and +5 oxidation states are clastogenic in vitro.f Various compounds.g Hexavalent chromium compounds only (e.g., K2Cr2O7, K2CrO4).h Nickel(II) salts (e.g., NiCl2) and insoluble crystalline nickel (e.g., Ni3S2).i Cobalt(II) salts only (e.g., CoCl2).j Cadmium(II) salts only (e.g., CdCl2).k A complex coal tar distillate that contains a variety of PAHs, PAH derivatives, and heterocyclic aromatic compounds [427].l Rodent dominant lethal assay only.m The TRI lists PACs (polycyclic aromatic compounds) as a category of 19 individual compounds. A list of compounds included is available
at http://www.epa.gov/tri/chemical/chemlist2001.pdf.n Primarily beryllium(II) compounds (e.g., BeSO4).o Drosophila SLRL (sex-linked recessive lethal) assay or aneuploidy assay only.p Animal cells only.q Classification restricted to chlorinated paraffins (alkanes) of average chain length C12 and average chlorination of approximately 60%.
Chromosomal effects (SCEs in vitro) observed for C12 compounds only.r Mixed isomers.s Drosophila sex-linked recessive lethal and mouse lymphoma assays only.t Also known as nitrofen.u Only 2,4-diaminotoluene evaluated.v 2B classification is for chlorophenoxy herbicides. 2,4-D, mecoprop (2-(4-chloro-2-methylphenoxy) propanoic acid), and methoxone ((4-
chloro-2-methyl-phenoxy) acetic acid) are classified as group 3 — inadequate evidence of animal carcinogenicity or inadequate evidence for a
definitive evaluation. Methoxone tests positive for mutagenicity in yeast, bacteria (Mutatox1 only), and Drosophila SLRL. Weak evidence of
SCE induction in vitro and in vivo in Chinese Hamsters and chick embryos.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345236
Table 3b
2002 NPRI on-site land releases of carcinogenic substances in Canada (>100 kg only)a
Compound Mutagenicity/clastogenicityb,c Carcinogenicityd Total land releases (�103 kg)
Lead compoundse CC 2B 1738.7
Asbestos (friable) C 1 521.2
Chromium compoundsf +++/CC 1 917.4
Nickel compoundsg ++/CC 1 265.6
Cadmium compoundsh ++/CC 1 150.8
Arsenic compoundsi +/CC 1 134.4
Cobalt compoundsj ++/CC 2B 42.6
Polycyclic aromatic compoundsk +++/CC 2A/2B 9.1
1,4-Dioxane +/C 2B 6.1
Ethylbenzene +l 2B 1.0
Benzene +++/CC 1 0.9
Chloroform +/CC 2B 0.5
Total inorganic 3770.7
Total organic 17.6
Grand total 3788.3
a Includes on-site landfill disposal, land treatment, surface impoundment, leaks, and spills. See Table 2b for additional details. Five other
organic compounds with values >10 and <100 kg (i.e., 1,2-dichloroethane, formaldehyde, acetaldehyde and vinyl acetate) account for an
additional 102 kg.b Based on data from the Genetic Activity Profile (GAP) database [454,455], the Monograph Series of the International Agency for Research
on Cancer (IARC) (available online at http://monographs.iarc.fr/), and the published results of the US national Toxicology Program (available
online at http://ntp-server.niehs.nih.gov/).c (�) Compounds for which there is no evidence of mutagenicity or clastogenicity, (+) mutagenic in bacterial and/or fungal cells in vitro, (++)
also mutagenic in plants or animal cells in vitro, (+++) also mutagenic in the Drosophila melanogaster somatic mutation and recombination test,
and/or sex-linked recessive lethal test, and/or transgenic rodent assays, and/or rodent dominant lethal assay. For cytogenetic endpoints C refers to
substances are clastogenic in in vitro assays, CC refers to substances that are clastogenic both in vitro and in vivo. Note: In some instances
conflicting results have been reported in the literature.d IARC classification system: (1) carcinogenic to humans, (2A) probably carcinogenic to humans, (2B) possibly carcinogenic to humans, (3)
inadequate or limited evidence of carcinogenicity in experimental animals.e Various compounds.f Hexavalent chromium compounds only (e.g., K2Cr2O7, K2CrO4).g Nickel(II) salts (e.g., NiCl2) and insoluble crystalline nickel (e.g., Ni3S2).h Cadmium(II) salts only (e.g., CdCl2).i Both the +3 and +5 oxidation states are clastogenic in vitro.j Cobalt(II) salts only (e.g., CoCl2).k Facilities had the option of reporting quantities for each of 17 individual PAHs, or, under a separate listing, the total for the 17 PAHs
categorized as toxic under section 11 of the Canadian Environmental Protection Act (1999, http://laws.justice.gc.ca/en/C-15.31/). Value is the
sum of IARC 2A carcinogens, IARC 2B carcinogens, and total PAH listings.l Animal cells only.
pollutants. For example, fine clay particles possess a
net negative charge and are capable of retaining
positively charged ions of toxic metals such as zinc,
copper, aluminum, chromium, arsenic, nickel, and
cadmium. Moreover, the oxidation state of a given soil
can dramatically modify the toxicity of metals such as
aluminum and arsenic.
Organic pollutants with limited water solubility
(e.g., aromatic hydrocarbons, chlorinated pesticides,
etc.) are usually sorbed to soil organic matter and clay
particles. The degree of sorption is primarily a
function of the solute’s water solubility and the soil
organic matter content, with solute vapour pressure
and soil clay content playing lesser roles. Sorption
potential of a given compound to a selected soil is
usually expressed as Kd, the slope of a linear sorption
isotherm [8]. Kd values that are corrected for organic
matter content are generally referred to as Kom or Koc,
with the subscripts referring to organic matter and
organic carbon, respectively [5].
The sorption of organic pollutants, a phenomenon
that is highly dependant on Kow (octanol–water
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 237
partition coefficient), is an important mechanism for
removal of pollutants from soil water, and consequent
inhibition of leaching to groundwater [4]. For
example, the mobility potential of most substances
with Kow values above 1000 are generally low to nil
[5]. Moreover, sorption of organic pollutants can alter
toxicity by reducing bioavailability and exposure.
Detailed discussions on the partitioning dynamics of
toxic organic pollutants in contaminated soils can be
found in Donnelly et al. [4], Winegardner [5], Young
et al. [9], Dragun [7], and Yaron et al. [10].
3.2. Detection of mutagens in soil
The spatial heterogeneity of the soil environment
complicates the selection of sampling locations, as
well as the selection of an appropriate sampling
method. Since budgetary considerations do not permit
the collection of an unlimited numbers of samples, a
variety of strategies and instruments have been created
to assist efficient sample collection and site assess-
ment. Three sample collection strategies are com-
monly employed to effectively characterize a site:
biased or likely location sampling, grid or pattern
sampling, and stratified random sampling [5,11]. Each
of these strategies offers a variety of advantages and
disadvantages. Biased sampling is simple and quick,
but generally reserved for situations of obvious
contamination. Prior site investigation or simple
visual surveys reveal obvious or likely areas of
contamination and these are selectively sampled (e.g.,
storage facilities, area containing damaged storage
drums). As such, this strategy is effective for
confirming site contamination, but not effective for
comprehensive risk assessment. Grid sampling,
collection of samples at regular intervals in a gridded
pattern that divides the site into an equal number of
uniform shapes, is easy to design and implement, and
provides excellent site coverage for identifying spills
and hot-spots. However, it is not practical for large
areas and may require an unacceptably large number
of samples. Stratified random sampling, a method that
is more cost effective than grid sampling, involves
collection of samples from a series of non-overlapping
strata that are defined on the basis of topological or
geophysical features. Random samples are collected
from each stratum, and because the variance within a
stratum is smaller than that between strata, the
combined results from numerous strata can provide
more accurate averages for selected parameters than
either grid or non-stratified random sampling [12–14].
However, this method requires competent topogra-
phical and/or geophysical surveys of the site prior to
sample collection. A detailed discussion of the
various soil collection strategies and the statistical
considerations of effective site monitoring are clearly
beyond the scope of this work. Interested readers
should consult Gilbert [12], Mason [13] and Cochran
[14].
Appropriate collection, handling, and storage of
soil samples depend on the depth required and the type
of analyses to be conducted. Soil samples are usually
collected with a surface sampler such as a spade or
scoop, a soil boring devise such as a manual or power
auger, or a soil coring devise that can provide an
undisturbed profile of soil condition and contamina-
tion. Scoop or spade sampling is the simplest and most
direct collection method that can readily provide small
quantities of surface cover to a specified depth. This
method is effective where disturbed shallow samples
are acceptable. Soil boring devises are commonly
used for subsurface sampling of disturbed soils from
depths below 20 ft. These devises (e.g., barrel auger,
continuous flight auger, etc.) permit continuous
removal of soil that can be sampled as it arrives on
the surface. Undisturbed soil samples are generally
collected using coring devises that push tube samplers
below the bottom of a borehole, or, in the case of the
hollow stem auger, the auger remains in place to
support the borehole walls while the core sampler is
inserted. More recent improvements on the hollow-
stem auger permit continuous coring as the auger
advances to the desired depth. Detailed descriptions of
soil collection techniques can be found in Wine-
gardner [5] and Boulding [15]. The latter publication,
an expanded version of the US EPA’s guide to the
description and sampling of contaminated soil [16],
also recommends a standard protocol for site and
sample description. Site descriptions should include
climate and weather (e.g., air temperature, humidity,
wind speed and direction), slope, vegetation, surface
erosion, and surface runoff. Sample descriptions
should include soil texture (i.e., sand, clay, silt
content), soil porosity, water content, and soil colour.
Detailed standardized methodologies can be found in
several ASTM publications [17,18].
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345238
Assessing the toxic contamination of a solid
medium like soil is not a simple task. Some of the
difficulties are not unique to soils and are common in
investigations of complex environmental samples such
as surface waters, groundwater, industrial effluents,
sediments, and airborne particulate matter. First, the
large number of toxic chemicals that may potentially
be present at a contaminated site can hinder successful
chemical analyses. Second, detailed chemical analysis
is limited in its ability to predict the toxicity of
complex chemical mixtures [19,20]. To overcome
these problems many researchers advocate a complex
mixture approach that employs bioassays to measure
the mutagenic potential of a complex environmental
sample or the extract/concentrate of an environmental
sample (e.g., for details see [19,21–29]). This
approach alleviates the need for chemical-specific
analyses, and integrates the effects of all mixture
components, whether or not they are known and
identified. Moreover, bioassay-directed chemical
analyses can subsequently be employed in an effort
to identify the putative mutagens (e.g., [22,23,30–
32]).
Soil texture, spatial heterogeneity, and microbial
content can further complicate assessment of muta-
genic potential. The most popular mutagenicity
assays, such as the Salmonella mutagenicity test
[33–36], require sterile samples and do not readily
accept solids. In addition, the low concentration of
mutagens in many environmental samples necessitates
extraction and concentration of substances prior to
mutagenicity assessment. Since a variety of known
environmental mutagens are organic compounds (e.g.,
PAHs, heterocyclic compounds, aromatic amines,
etc.) [37], many researchers employ organic solvents
to extract and concentrate organic constituents prior to
mutagenicity testing.
Dichloromethane (DCM) and methanol (MetOH),
as well as solvent mixtures such as acetone/hexane,
are the most popular extraction solvents, and many
recent publications investigating the mutagenic
hazards of contaminated soils examined the muta-
genicity of DCM, MetOH or acetone/hexane extracts
of dried soils (e.g., [38–57]). Other extraction solvents
or solvent mixtures include acetonitrile (ACN) [58–
60], acetone [60,61], ethyl ether [62], acetone/
cyclohexane [63], DCM/MetOH [64], and ethanol/
dimethyl sulfoxide [65]. Some researchers employing
plant assays for soil mutagenicity assessment do not
expose plant tissues (e.g., cuttings, root tips, etc.) to
organic soil extracts [66–69]. Rather, plant tissues are
exposed directly to unaltered soil [70–80], either in
situ or ex situ, or tissues are exposed to aqueous
extracts, leachates, or elutriates [63,67–69,76,81–85].
Although many researchers have employed bioas-
says to investigate the sources and identity of
mutagens and carcinogens in complex samples from
aquatic systems (e.g., [19,86,87]), there is a paucity of
information about the sources and potential hazards of
mutagens in soil. Moreover, few researchers have
investigated the efficacy of short-term mutagenicity
assays such as the Salmonella mutagenicity test in the
assessment of soil mutagenic hazard. Thus, there is a
need to investigate the mutagenic hazards of
contaminated soils, and moreover, incorporate the
use of well-established assays in assessments of
potential hazard. This work provides a comprehensive
review of published research that employed bioassays
to investigate the mutagenic properties of soil.
3.3. Bioassays employed for soil mutagenicity
assessment
Published accounts of soil genotoxicity assessment
have employed more than 30 assays to assess DNA
damaging ability, mutagenicity, or clastogenicity.
These include popular bacterial and plant assays such
as the Salmonella mutagenicity test, the anaphase
aberration test in Allium cepa and the Tradescantia
micronucleus test, as well less common tests such as
micronucleus (MN) induction test in Xenopus laevis
[88,89] and rifampicin resistance forward mutation
test in Pseudomonas putida [90]. Fig. 1 provides a
breakdown of the genotoxicity assessments techniques
employed in published examinations of contaminated
soils. The figure shows that 37.6% of the published
assessments employed the Salmonella mutagenicity
test; with a further 13.4% employing other prokaryote
mutagenicity and DNA damage tests (e.g., SOS
Chromotest, Mutatox1, l prophage induction assay,
Salmonella umu test, rec� differential survival assay).
The popularity of the Salmonella mutagenicity test for
the assessment of soil mutagenicity is similar, but
notably lower, than that described by Houk [19] for
assessments of industrial wastes and effluents. Houk
[19] indicated that almost 60% of works that
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 239
Fig. 1. Breakdown of the assays employed in published assessments of soil genotoxicity.
investigated the mutagenicity of industrial effluents
and wastes employed the Salmonella mutagenicity
test. Most of the remaining assessments (i.e., 37.6% of
the total) employed one of several plant mutagenicity
or clastogenicity assays (e.g., anaphase aberrations
test in Allium sp., stamen hair mutation test in
Tradescantia sp.). The remaining 11.6% of the
assessments employed a variety of assays examining
cytogenetic effects (e.g., sister chromatid exchanges
or SCEs) in cultured human lymphocytes, somatic
mutations in Drosophila, forward mutations in
Aspergillus nidulans, and hprt mutations in Chinese
Hamster V79 cells.
Fig. 2 provides a breakdown of the genotoxicity
endpoints employed in published examinations of
contaminated soils. The majority (i.e., 60%) of the
assessments measured induction of gene mutations.
Loci examined include the his loci in Salmonella
[33,34,36], the stamen hair colour loci in Tradescan-
tia [91], the waxy locus in Zea mays [72], the mwh and
flr wing morphology loci in Drosophila melanogaster
[92], the hprt locus in Chinese Hamster V79 cells
[93], the methionine suppressor loci in Aspergillus
nidulans [94,95], and the ade loci of Saccharomyces
cerevisiae [96]. Twenty-seven percent of published
soil genotoxicity assessments employed a variety of
clastogenicity endpoints such as chromosomal aber-
rations (CAs), MN induction, and the induction of
SCEs. Although enhanced risk of gene mutation is
associated with a variety of adverse health effects
including cancer (e.g., [97–100]), only two of the
clastogenicity endpoints (e.g., MNs and CAs) are
known to be useful biomarkers of adverse health
effects such as cancer [101–103]. SCEs are useful
markers of cytogenetic effects in vitro and in vivo
[104,105]; however, their utility as biomarkers of
adverse health effect is disputed [102,106]. Much of
the remaining 12.7% of genotoxicity assessments
measured induction of DNA damage using assays
such as the rec� differential survival assay in Bacillus
subtilus [107,108], the SOS Chromotest [109,110],
the Salmonella umu test [111], and the Microscreen
prophage induction assay [112].
4. Collection and analysis of published soilmutagenicity/clastogenicity data
In total, 165 published assessments (i.e., endpoint/
sample combinations) of soil genotoxicity were
obtained from 118 publications, including thirteen
foreign language publications (e.g., Spanish, Russian,
Italian and Japanese). These included journal articles,
book chapters, government reports, and conference
abstracts.
For the purposes of empirical analyses, the
reviewed soil genotoxicity assessments were divided
into two main groups. Those that used popular assays
for which large amounts of published data are
available (i.e., >30 observations), and those that
employed assays rarely used for soil genotoxicity
assessment. Five assay systems have been frequently
used for soil genotoxicity assessment, the Salmonella
mutagenicity test and four plant assay systems: the
stamen hair mutation assay in Tradescantia, the
Tradescantia MN assay, the anaphase aberration assay
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345240
Fig. 2. Breakdown of endpoints employed in published genotoxicity assessments of soils and soil extracts. Gene mutation endpoints include his
loci in Salmonella, the Wx locus in Zea mays, the hprt locus in mammalian cells, stamen hair colour loci in Tradescantia, and methionine
suppressor loci in Aspergillus. Chromosome damage endpoints include chromosome aberrations, micronucleus formation, sister chromatid
exchanges, mitotic recombination, gene conversion, and aneuploidy. DNA damage endpoints include SOS response induction (e.g., rec
dependant DNA repair, prophage induction, sulA induction, umu induction), formation of DNA adducts, and induction of alkali-labile DNA
damage sites. Miscellaneous endpoints include the formation of nuclear anomalies in maize and white clover (Trifolium repens) assessed via flow
cytometry.
in Allium cepa root tip, and the waxy locus mutation
assay in Zea mays.
Soil contamination data (e.g., PAHs, dinitropyr-
enes, 137Cs, etc.) were also collected where available.
All values were converted to a common unit such as
ppm dry weight (i.e., mg/g dry soil) or Bq per dry kg.
PAH contamination values were collected from 13
studies that employed the Salmonella mutagenicity
assay, four studies that employed the Tradescantia
MN assay, and one study that assessed induction of
rifampicin resistance mutations in Pseudomonas
putida. Soil PAH data for one Salmonella study
[39] were obtained from a separate, but related
publication [113]. Unfortunately, there was little
consistency across the studies with respect to the
identity of the PAHs examined. Three studies provided
total PAH values, but failed to provide a list of
measured PAHs [57,114,115]. Seven studies provided
data for 10 to 14 PAHs, including several noteworthy
mutagenic carcinogens such as benzo[a]pyrene,
benz[a]anthracene, dibenz[ah]anthracene, and ben-
zo[b]fluoranthene [39,59,63,113,116–118]. Six stu-
dies provided data for 3–8 PAHs selected to indicate
anthropogenic contamination by pyrolytic emissions
(e.g., fluoranthene, chrysene, pyrene, benzo[a]pyrene)
[56,83,119–122]. Despite these variations in the
identity of the PAHs selected for analysis, all the
selected PAHs are members of the list of 16 PAHs
commonly referred to as priority PAHs [123].
For studies that employed the Salmonella muta-
genicity test, total PAH was operationally defined as
the total PAH value provided by the author(s), or
the sum of the most frequently measured PAHs:
fluoranthene, pyrene, benz[a]anthracene, chrysene,
benzo[a]pyrene, benzo[b]fluoranthene, benzo[k]fluor-
anthene, and benzo[ghi]perylene. A matrix of Pearson
correlations between eight measured PAHs showed
high correlations between individual PAH concentra-
tions across the studies examined, with an average
coefficient of 0.92 (0.81–0.99). Thus, where the
concentration of one or more of the required PAHs was
not available, predictive regression models based on
all available data were used to predict missing values.
With respect to the Tradescantia studies, Total PAH
simply refers to the sum of individual values available
from the relevant publications.
Four studies provided detailed information on a
series of dinitropyrenes (DNPs) [50,51,53,124]. For
the purposes of data analyses, Total DNPs was
operationally defined as the sum of 1,3-dinitropyrene,
1,6-dinitropyrene, and 1,8-dintropyrene.
Several studies that employed the Tradescantia
MN assay also measured soil concentrations of several
metals (e.g., As, Cd, Sb, Cr, Cu, Ni, Pb, Zn)
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 241
Table 4
Overview of the collected soil contamination dataa
Assay system Analyte Minimum Maximum Geometric mean N
Salmonella TA98 with S9 PAHs 0.07 12266 11.0 � 4.7 80
Salmonella TA98 without S9 DNPs 0.03 15.7 0.62 � 0.15 37
Tradescantia MN assay PAHs 0.02 5749 2.6 � 2.4 17
Cr 0.05 214 7.7 � 1.6 34
Pb 0.08 28144 101.0 � 48.8 34
As 0.02 2940 7.2 � 3.7 34
Cd 0.01 89.9 0.7 � 0.3 34
Sb 0.2 6720 2.9 � 2.6 34
Cu 0.03 3203 13.5 � 7.0 34
Ni 0.08 366 10.5 � 2.84 34
Zn 0.02 23727 66.1 � 26.9 34
Allium cepa CA assay 137Cs 22.0 6880 2084.1 � 354.2 17
Arabidopsis mutation 137Cs 22.0 2529 333.5 � 200.4 4
a Values expressed as ppm dry wt. (i.e., mg/kg), except DNPs, which are ppb dry wt. (i.e., mg/kg), and 137Cs, which are Bq/kg. All metal
concentrations are total extractable metals.
[63,76,77,81,83]. However, total concentrations of Cr,
Pb, As, Cd, Sb, Cu, Ni, and Zn were only collected
from two studies that conducted direct soil exposures
of intact Tradescantia plants [76,77]. Table 4 provides
an overview of the soil contamination data that was
used to investigate empirical relationships between
bioassay response and soil contamination.
All data analyses were performed using the SAS
system version 8.02 for Windows [125]. Analysis of
variance (ANOVA) was employed to investigate
relationships between bioassay response (e.g., Sal-
monella revertents/mg dry weight) and site classifica-
tion (e.g., urban/industrial, remote). Ordinary least-
squares linear regression analysis was used to
investigate relationships between genotoxic potency
and measures of environmental contamination (e.g.,
PAH concentration). In some cases, analysis of
covariance (ANCOVA) was employed to simulta-
neously investigate the effects of site classification and
site contamination. Following the notation of Gujarati
[126]], the general model Yi = a1 + a2(D2) + a3(D3)
+ an(Dn) + b1(Xi) + ((b2 � D2) � (Xi)) + ((b3 � D3)
� (Xi)) + ((bn � Dn) � (Xi)) + mi was fit to the data. Yi
is the observed mutagenic potency value, Xi is the
value for a continuous environmental contamination
variable at observation i (e.g., PAH concentration,
where available), and D2 through Dn are dichotomous
variables that indicate membership of observation i in
a given group (e.g., remote sites, urban/industrial sites,
etc.). D2 through Dn are set to 1 when the condition of
group membership is satisfied and 0 when the
condition is not satisfied. This model permits an
assessment of the relationships between genotoxic
potency and a multitude of continuous or dichotomous
variables taken individually or in various combina-
tions. The scalars D2 through Dn permit adjustments in
slope and intercept values where appropriate. The
residual error term mi was assumed to be independent
and normally distributed and normality was assessed
using the Shapiro-Wilk test and visual examination of
a normal probability plot [127]. Where necessary the
data were log transformed to meet the assumptions of
least squares regression, ANOVA and ANCOVA. The
absolute value of the residual error values was used to
detect outliers and identify data entry errors. To
identify significant outliers, externally studentized
residuals (di*) were calculated for each residual [128].
Comparison of di* values to the appropriate t
distribution permitted the identification of significant
residuals, which lie beyond the 0.05 limits of the
distribution.
5. Salmonella mutagenicity of soils
The Salmonella mutagenicity test is undoubtedly
the most popular bioassay in environmental mutagen-
esis research, particularly for the analysis of complex
mixtures such as organic extracts of soil, air, and water
[26–29,31,129–133]. The standard version of the
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345242
assay, known as the plate incorporation assay, is a
reverse mutation test that quantifies the frequency of
reversion from histidine auxotroph to wild-type
following a 48- to 72-h incubation with the test
substances [33–36]. Several tester strains of Salmo-
nella are available, carrying a variety of his mutations.
The most popular tester strains, TA98 and TA100,
carry the hisD3052 and hisG46 alleles, respectively.
The former is a �1 frameshift mutation reverted to
wild type by frameshift mutagens (e.g., ICR-191,
nitrosamines). The latter carries a base-substitution
mutation that is reverted by base-pair substitutions at a
GC pair in a proline codon [33]. These strains have
been extensively employed for the detection of
environmental mutagens including PAHs [134–136],
nitroarenes [137,138], aromatic amines (e.g., N-
containing heterocyclics) [139–141], S-containing
heterocyclics [142,143], and phenylbenzotriazoles
[144,145]. Although some of these compounds have
noteworthy mutagenic activity on both TA100 and
TA98 (e.g., PAHs) [134], several are known to have
more potent frameshift activity (e.g., aromatic amines,
phenybenzotriazoles, nitroarenes) [137,139,141,144].
Sixty-two published works contained Salmonella
mutagenicity data on soil extracts. The majority of
these studies employed the standard plate incorpora-
tion version of the assay with Salmonella strains TA98
and/or TA100. Where mutagenic potency values were
not provided and the published data showed at least a
two-fold increase in response over the control,
ordinary least squares regression was employed to
calculate potency from the initial (linear) portion of
the concentration–response relationship. In all cases,
reported mutagenic potency values were converted to
net Salmonella revertents per equivalent dry mg of
soil. Some researchers examining soil extracts have
referred to potency expressed as net revertents per g
dry soil as weighted activity [56,114,115,117,118].
In several cases, missing information (e.g., yield of
extractable material per gram of soil) or additional
details about sites and/or extraction procedures were
obtained directly from the corresponding author
[53,56,146,147]. If mutagenic potency was reported
for individual chemical fractions (e.g., base/neutral
compounds), total mutagenic potency was defined as
the sum of the individual fractions. In cases where the
number of spontaneous revertents was not reported,
the following values were used: 25 for TA98 without
S9, 35 for TA98 with S9, 100 for TA100 without S9,
and 120 for TA100 with S9. Salmonella mutagenicity
data were not collected from studies that only
provided a qualitative response index (e.g., positive/
negative) [147,148], studies that did not provide
sufficient information about concentration [149,150],
or sufficient information to calculate mutagenic
potency in revertents/mg dry soil [38,44,62,151–
155]. The majority of the studies in the latter category
expressed mutagenic potency as net revertents per unit
of extractable organic material (e.g., mg extractable
residue or mL of solvent extract), and did not provide
the information required (i.e., mg extractable material
per mg dry wt. of soil) to convert the potency to
revertents per gram of soil.
A total of 1633 observations of Salmonella
mutagenic potency were collected from the literature.
Over 50% of these values were generated from tests of
MetOH extracts. Roughly 24% of the values were
generated from examinations of hexane/acetone
extracts, and 14% were generated from DCM extracts.
The remaining 11% of the mutagenic potency values
were generated from examinations of extracts that
employed a variety of solvents including ACN,
cyclohexane, or solvent mixtures (e.g., hexane/
isopropanol, DCM/MetOH, benzene/EtOH, etc.).
Appendix A contains the Salmonella mutagenicity
data obtained from the literature.
Site descriptions in the publications, or information
obtained from the corresponding author, were used to
divide the Salmonella data into three separate site
categories: rural/agricultural, urban/suburban, and
industrial. Rural/agricultural sites are all located
outside urban or suburban communities and include
remote park-like settings or agricultural areas. Urban/
suburban sites include sites located in cities or
towns but not directly on the grounds of an industrial
facility that receives industrial wastes and discharges.
Category assignments were based on site descriptions
only, without prior examination of the mutagenicity
data. Table 5 shows the distribution of the data
between the three site categories.
5.1. TA98 and TA100 mutagenicity of soil extracts
Much of the soil mutagenicity literature is
concerned with the identification of hazardous sites
that may pose a mutagenic hazard to humans and
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 243
Table 5
Breakdown of the Salmonella mutagenicity data collected from the
literature
Site/sample category Number of observations
Salmonella strain Total
TA98 TA100
No S9 +S9 No S9 +S9
Rural/agricultural 125 144 92 109 470
Urban/residential 219 241 157 199 816
Industrial 105 202 15 25 347
Total 449 587 264 333 1633
indigenous fauna (e.g., [44,45,47,117,118]), and the
predominance of particular Salmonella strain-activa-
tion combinations in the literature reflects their utility
in this regard. Much of the published Salmonella
mutagenicity data on soil extracts are TA98 results,
often obtained in the presence of an S9 metabolic
activation mixture (see Table 5). This suggests that
much of the mutagenic activity in contaminated soils
is S9-activated frameshift activity. This assertion is
supported by numerous studies of organic extracts
from soils contaminated with a wide range of
pollutants (e.g., wood preserving wastes, petrochem-
ical wastes, sewage sludge) that yielded little or no
response on the base-substitution strain TA100 (e.g.,
[38,41,47,48,54,64,115,116,156]). Moreover, many of
these studies have noted the presence of known
frameshift mutagens such as nitroarenes and N-
containing heterocyclics in soil extracts that show
potent TA98 activity [64,116,121,157]. However, it
should be noted that some studies, such as those that
examined extracts of soils contaminated with muni-
tions wastes (e.g., di- and trinitrotoluenes) and
dintropyrenes (e.g., 1,3-, 1,6-, and 1,8-dinitropyrene),
also showed strong responses in Salmonella TA100
(e.g., [51,158]).
Table 6 provides a descriptive summary of the
Salmonella mutagenic potency values for each
combination of site category and Salmonella strain.
The data presented illustrate that mutagenic potency
values range from a low of less than 0.001 revertents
per dry mg at rural sites, to a high of almost 1000
revertents/mg at highly contaminated industrial sites.
Fig. 3 contains a box plot that illustrates the
distribution of the log transformed Salmonella
TA98 results obtained in the presence of S9 metabolic
activation (N = 587). Analysis of variance (ANOVA),
employed to investigate the relationship between
mutagenic potency and site category, revealed a
significant relationship between site category and
mutagenic potency (p < 0.0001), and a significant
difference between rural, urban, and industrial sites
(geometric mean values = 0.06, 0.47, and 0.95
revertents/mg, respectively).
The box plot for industrial soils shown in Fig. 3 (top
left) suggests that several observations would likely be
considered to be outliers. Studentized deleted resi-
duals (di*) for each observation were calculated to
identify significant outliers from the ANOVA model.
Based on di* values, 40 of the mutagenic potency
values shown in Fig. 3 (top left) were identified as
significant outliers. Thirty-nine of these outliers are
industrial sites, with almost half of them (43%) being
negative outliers with potency values below 0.02
revertents/mg. These results are not surprising in light
of the fact that some low potency values were assigned
to the industrial category based solely on the published
site descriptions, and many of the negative outliers
reflect results obtained for samples collected on
transects extending a considerable distance from the
source(s) of the industrial contamination (e.g., coal
gasification site sample 201 from Donnelly et al. [118]
and munitions site soil 006 from Donnelly et al. [54]).
The other negative outlier is a sample from an urban
site in Azerbaijan. The authors described the sample
collection area as a location ‘‘near apartments in a
highly populated urban area’’ [56].
The remaining significant industrial outliers (i.e.,
57%) are positive outliers with potency values above
55 revertents/mg. All of these positive outliers, with
extremely high mutagenic potency values (i.e., 56.9–
376 revertents/mg), are from heavily contaminated
sites such as the Superfund site studied by McDaniels
et al. [46], the tar pit perimeter studied by Donnelly et
al. [118], the explosive contaminated soils studied by
Donnelly et al. [54], Griest et al. [158], and Berthe-
Corti et al. [65], and the petrochemical and wood
preserving wastes studied by Brown et al. [49].
Fig. 3 (top right) contains a box plot that illustrates
the distribution of the log transformed Salmonella
TA98 results obtained in the absence of S9 metabolic
activation (N = 449). The ANOVA results reveal a
significant relationship between site category and
mutagenic potency (p < 0.0001), and a significant
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345244
Table 6
Descriptive summary of the Salmonella mutagenicity dataa
Category
Rural Urban/suburban Industrial
TA98 with metabolic activation (N = 587)
N 144 241 202
Minimum 0.0020 0.0083 0.00070
Maximum 0.77 10.90 376.00
Mean 0.10 1.04 21.67
S.E.b 0.0091 0.11 3.81
Geometric meanc 0.060 0.47 0.95
Distributiond No No No
TA98 without metabolic activation (N = 449)
N 125 219 105
Minimum 0.00029 0.032 0.0020
Maximum 0.50 46.8 288.35
Mean 0.095 1.35 24.69
S.E. 0.0092 0.30 5.91
Geometric mean 0.057 0.43 0.77
Distribution No No No
TA100 with metabolic activation (N = 333)
N 109 199 25
Minimum 0.00078 0.047 0.016
Maximum 1.04 6.72 925.11
Mean 0.15 0.75 93.72
S.E. 0.016 0.067 43.18
Geometric mean 0.096 0.45 3.18
Distribution No Yes Yes
TA100 without metabolic activation (N = 262)
N 92 157 15
Minimum 0.0019 0.029 0.0075
Maximum 0.64 4.89 259.00
Mean 0.17 0.39 17.77
S.E. 0.014 0.044 17.24
Geometric mean 0.12 0.26 0.13
Distribution No Yes NAe
a Data obtained using standard plate incorporation version of the Salmonella mutagenicity test on organic soil extracts. All values are
mutagenic potency values expressed as Salmonella revertents per equivalent mg dry soil.b Standard error of the mean.c Geometric mean.d Result of Shapiro-Wilk test for normality on log-transformed values. No indicates that the null hypothesis of normal distribution was
rejected at p < 0.05.e Insufficient data for normality test.
difference between rural, urban, and industrial sites
(geometric mean = 0.06, 0.43, and 0.77 revertents/mg,
respectively). The di* values revealed 37 significant
outliers, 31 of which are industrial sites. More than a
third (39%) of these industrial sites yielded organic
extracts with unexpectedly low potency values (i.e.,
below 0.03 revertents/mg). The remaining industrial
outliers (i.e., 61%) are those that yielded extremely
potent extracts (e.g., above 30 revertents/mg). Eigh-
teen of the 19 positive outliers are soils contaminated
with munitions and explosive wastes studied by
Donnelly et al. [54], Berthe-Corti et al. [65], and
Griest et al. [158]. This may not be surprising since di-
and trinitrotoluenes and related compounds found in
munitions contaminated soils (e.g., HMX and RDX)
are known to be potent direct-acting frameshift
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 245
Fig. 3. Box plots of published Salmonella (TA98 and TA100) mutagenic potency values for organic extracts of soils collected from heavily
contaminated, urban/industrial, and remote sites. From bottom to top, the horizontal solid lines in each box represent the 10th, 25th, 50th, 75th,
and 90th percentiles of the distributions. The dotted horizontal line is the mean value. Suspected outliers are shown as dots. The overlayed text
shows the results of the ANOVA analysis for site category effect. Boxes labeled with different letters are significantly different at p < 0.05
(Duncan multiple range test [127]). Potency values were log transformed to meet the assumptions of least squares ANOVA. The number of
observations in each site category is available in Table 5. The sources of all data are shown in Appendix A.
mutagens [159,160]. The remaining positive outlier is
an extract of a soil sample experimentally treated with
diesel oil [157]. Diesel fuel, a petroleum middle
distillate composed of a complex mixture of alkanes,
cycloalkanes and aromatic compounds in the C9 to C28
range, can contain high concentrations (5–10 wt.%) of
3- to 7-ring aromatic compounds [161]. However, it
should be noted that several separate studies noted that
the Salmonella mutagenicity of diesel oil was found to
be weak or equivocal [161–164], and it is not
immediately obvious what components in a diesel
oil treated soil could be responsible for a potent direct-
acting response on TA98.
Three rural sites yielded extremely low potency
values (e.g., �0.001 revertents/mg) that were sig-
nificant negative outliers. These are reference soils
used in numerous studies of contaminated soils,
amended soils, and remediated soils conducted by
researchers at Texas A&M University [40,42,48,
49,165,166]. The three remaining outliers are urban
sites in Japan that yielded highly potent samples and
large positive residuals (e.g., Hekinan). These sites are
known to be contaminated with combustion related
dinitropyrenes (e.g. 1,3- 1,6- and 1,8-dinitropyrenes)
that account for 18–51% of the observed direct acting
TA98 mutagenicity [52,53].
Fig. 3 (bottom left) contains a box plot that
illustrates the distribution of the log transformed
Salmonella TA100 results obtained in the presence of
S9 metabolic activation (N = 333). The ANOVA
revealed a significant relationship between site
category and mutagenic potency (p < 0.0001), and
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345246
Table 7
Geometric mean Salmonella mutagenicity valuesa
Category
Rural Urban/suburban Industrial
TA98 with S9 0.06 � 0.005 0.47 �0.05 0.95 � 0.17
TA98 without S9 0.057 � 0.006 0.43 � 0.10 0.77 � 0.18
TA100 with S9 0.096 � 0.01 0.46 � 0.04 3.18 � 1.46
TA100 without S9 0.12 � 0.01 0.26 � 0.03 0.13 � 0.13
a Salmonella revertents per equivalent mg dry soil � standard
error.
a significant difference between rural, urban, and
industrial sites (geometric mean = 0.096, 0.46, and 3.2
revertents/mg, respectively). The di* values revealed
15 significant outliers including eleven industrial, two
urban, and two rural/agricultural sites. Five of the
industrial outliers as well as the two rural outliers
yielded low potency values and were significant
negative outliers. The two rural outliers are the
aforementioned reference soils used by researchers at
Texas A&M University. Four of the five negative
outliers from industrial sites are samples collected in
the vicinity of a carbon electrode production facility.
The authors of the study [63] describe the area as ‘‘a
contaminated area in Lombardy, near a factory for
carbon electrode production’’ and the sampling site as
‘‘highly contaminated’’ with PAHs. Nevertheless,
cyclohexane/acetone extracts from the collected
samples yielded low mutagenic potency values. The
fifth outlier is a weak extract of an agricultural soil
treated with industrial and domestic wastewaters [60].
The remaining eight outliers were all significant
positive outliers, six industrial and two urban. The two
urban samples were collected from highly urban sites
located in the Aishi and Hokkaido prefectures of Japan
(e.g., Hekinan, Muroran) [53]. More specifically,
regions previously recognized for contamination by
atmospheric mutagens (e.g., PAHs, DNPs) from
gasoline and diesel powered vehicles [50–52]. The
six industrial outliers yielded potency values between
87 and 925 revertents/mg associated with extracts of
six soils contaminated by petrochemical or wood
preserving wastes [49], and one extract of an explosive
contaminated soil [158].
Fig. 3 (bottom right) contains a box plot that
illustrates the distribution of the log transformed
Salmonella TA100 results obtained in the absence of
S9 metabolic activation (N = 264). The ANOVA
reveals a significant relationship between site category
and mutagenic potency (p < 0.0001), and a significant
difference between rural and urban sites, and urban
and industrial sites (geometric mean = 0.12, 0.26, and
0.13 revertents/mg for rural, urban, and industrial
sites). However, despite industrial potency values
greater than 200 revertents/mg for an explosive
contaminated soil [158], the results did not reveal a
significant difference between rural and industrial
sites. This is most likely caused by the paucity of
industrial data, combined with the large range and
skewed nature of the industrial data (e.g., 8 of the
industrial potency values are <0.01 revertents/mg).
The di* values revealed 12 significant outliers,
including 6 of the 13 industrial sites, 3 urban sites,
and 3 rural sites. All three rural sites yielded low
potency values (e.g., <0.01 revertents/mg) that were
significant negative outliers. All four of the industrial
sites that yielded negative outliers, were samples
collected in the vicinity of the aforementioned carbon
electrode production facility [63]. Both of the
industrial soils that yielded significant positive outliers
are soils contaminated with explosive residues or
munitions wastes that elicited potency values between
6 and 259 revertents/mg [65,158]. Again, these
elevated potency values are presumably related to
the potent direct-acting activity of munitions com-
pounds (e.g., TNT and related compounds) on both
TA98 and TA100 [159,160].
The three urban outliers include one negative
outlier from a public park in Japan [53], and the two
positive outliers are heavily urbanized sites in Japan
and Germany [51,53,116]. One of the latter sites is the
same positive outlier identified in the analyses of
TA100 S9-activated results (i.e., Hekinan, Aishi
Prefecture).
The geometric mean Salmonella mutagenicity
results for each site category (Table 7) indicate that
sites unimpacted by urban or industrial emissions may
be expected to yield positive Salmonella mutagenicity
responses with mean TA98 potency values of roughly
0.06 revertents/mg and mean TA100 potency values of
roughly 0.1 revertents/mg. These values are in
agreement with several ‘‘background’’ soil mutageni-
city values noted in the literature. Jones and Peace [39]
stated that the natural background level of Salmonella
mutagenicity in soil can be expected to be less than 0.1
revertents/mg. In their study of DCM extracts of
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 247
Welsh soils they observed 0.006–0.019 revertents/mg
for TA98 and 0.024–0.038 revertents/mg for TA100. A
study by Donnelly et al. [114] of an abandoned solvent
recovery site indicated that TA98 mutagenicity values
for ‘‘background’’ soils are less than 0.02 revertents/
mg (without S9). Jones and Page [167] further assert
that values in the 1.0 to 1.5 revertents/mg could be a
‘‘trigger’’ for identifying contaminated, potentially
hazardous soils that are worthy of concern. This value
is in agreement with the more rigorously determined
geometric mean values for urban/suburban and
industrial soils summarized in Table 7 (i.e., 0.29–
0.43 revertents/mg for urban sites and 0.81–4.0
revertents/mg for industrial sites).
5.2. Relationships between Salmonella mutagenicity
and extraction solvent
The abundance of TA98 mutagenicity data per-
mitted an examination of the effect of extraction
solvent on the mutagenic potency of soil extracts.
Two-way ANOVA was conducted on the direct-acting
data (N = 392) to investigate a site category effect, an
extraction solvent effect (DCM, MetOH, acetone/
hexane only), and a solvent-site interaction (i.e., an
extraction solvent effect that differs with respect to site
category). The results revealed a significant solvent
effect (p < 0.005) and a significant solvent-site
interaction (p < 0.005). Post-hoc examination of the
mean values for each ANOVA cell indicated that for
industrial soils DCM extracts are almost twice as
potent as MetOH extracts. Conversely, for rural/
agricultural soils, the results suggest that MetOH and
acetone/hexane extracts are an order of magnitude
more potent than DCM extracts. Two-way ANOVA of
the S9-activated values (N = 511) failed to reveal a
significant solvent (DCM, MetOH, acetone/hexane
only) effect, but revealed a significant (p < 0.0001)
solvent–site interaction. Post-hoc analyses of the
mean values indicated that DCM extracts of industrial
and urban soils are more than seven-fold more potent
than their respective MetOH extracts, but the
differences between mean values for rural/agricultural
soils are negligible.
Despite the fact that the TA100 data set is
considerably smaller than the TA98 data set, two-
way ANOVA was also used to investigate extraction
solvent effects (DCM, acetone/hexane, MetOH) and
solvent-site interactions (S9-activated only). The
results obtained (N = 306) revealed a marginal solvent
effect (p < 0.02), and a significant site–solvent
interaction (p < 0.0001). Again, post-hoc analyses
of mean values suggest that the S9-activated mutagens
extracted from industrial soils are considerably more
soluble in DCM. Comparison of mean values showed
that the average potency of DCM extracts from
industrial soils are more than 20-fold greater than
acetone/hexane extracts. Conversely, post-hoc ana-
lyses of the rural/agricultural data suggest that
mutagens in these soils are soluble in more polar
solvents such as MetOH. Examination of these mean
values showed that MetOH extracts are, on average,
more than two-fold more potent than DCM extracts.
Unfortunately, limitations in the data did not permit
more detailed analyses (i.e., study-specific) of the S9-
activated TA100 potency values or similar analyses for
the direct-acting TA100 data.
Although these results are interesting, and suggest
that organic mutagens in contaminated industrial soils
are lipophilic, the results are clearly biased by
unbalanced ANOVA cell sizes, and variability in
solvent choice across different studies examining very
different sites. For example, DeMarini et al. [41]
examined DCM extracts of PCB contaminated soil
and recorded TA98 (with S9) mutagenic potency
values of 0.1–0.2 reverants/mg dry soil. In contrast,
McDaniels et al. [46] examined cyclohexane extracts
of Superfund soils and recorded mutagenic potency
values that are two orders of magnitude greater than
those of DeMarini et al. Inference that cyclohexane
extracts are generally more potent than DCM extracts
would be misleading. A robust investigation of an
extraction solvent effect requires matched mutagenic
potency values (i.e., two or more extracts of sub-
samples of a single soil). Several studies (e.g., [46,47])
contain the data required for a balanced ANOVA
investigating an extraction solvent effect. The results
obtained, summarized in Table 8, indicate that only
four of the twelve studies examined reveal a
significant extraction solvent effect. These study-
specific results indicate that the less polar extraction
solvents (e.g., cyclohexane, DCM) tended to provide
more potent samples than the more polar solvents
(e.g., MetOH). This assertion is in agreement with the
aforementioned results based on the S9-activated
TA98 data. Additional analysis of industrial values
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345248
Table 8
Effect of soil extraction solvent on Salmonella TA98 mutagenicity
Data Extraction solvents N F-ratio Significance
TA98 without S9
All industrial sites DCM, MetOH 79 0.61 No
Abandoned solvent recovery site [114] DCM, MetOH 10 0.54 No
Munitions contaminated soils [54] DCM, MetOH 28 1.48 No
Abandoned chemical manufacturing site [47] DCM, MetOH 12 0.53 No
Soils amended with sewage sludge [156] DCM, MetOHa 12 5.54 Yes*
All rural/agricultural sites DCM, MetOH, Ac/Hexb 118 21.27 Yes***
TA98 with S9
All industrial sites DCM, MetOH, Cyclohex, Ac/Hexc 167 5.20 Yes**
Industrial sites (DCM, MetOH only) DCM, MetOHd 163 10.09 Yes**
Abandoned wood preserving facility [115] DCM, MetOHe 24 4.49 Yes*
Superfund sites (unspecified) [46] DCM, Cyclohexf 14 12.64 Yes**
Munitions contaminated soils [54] DCM, MetOH 28 0.14 No
Hazardous waste dump site [117] DCM, MetOH 12 0.23 No
Abandoned coal gasification site [118] DCM, MetOHg 28 8.45 Yes**
Abandoned chemical manufacturing site [47] DCM, MetOH 12 0.49 No
Soils amended with sewage sludge [156] DCM, MetOH 12 0.08 No
Hazardous waste landfill [423] DCM, Ac/Hex 6 0.21 No
All urban/residential sites DCM, Ac/Hex, MetOHh 214 47.18 Yes***
All rural/agricultural sites DCM, MetOH, Ac/Hexi 130 12.46 Yes***
a DCM > MetOH.b Ac/Hex > MetOH > DCM.c DCM > Ac/Hex > MetOH.d DCM > MetOH.e DCM > MetOH.f Cyclohex > DCM.g DCM > MetOH.h MetOH > DCM > Ac/Hex.i Ac/Hex/> MetOH > DCM.* p < 0.05.** p < 0.01.*** p < 0.001.
alone failed to reveal a significant solvent effect
without S9, but revealed a significant effect with S9.
The latter analysis indicated that less polar solvent
such as DCM are more effective at extracting TA98
mutagens that require S9 activation from soils
collected at industrial sites.
Although based on limited data, the results
presented in Table 8 also suggest that the more polar
solvents such as MetOH preferentially extract muta-
gens (TA98 with and without S9) from rural/
agricultural (i.e., non-industrial) soils. Again, this is
in agreement with the aforementioned analyses
conducted on the entire set of TA98 data. Although
the results also suggest that MetOH may be more
effective at extracting mutagens (TA98 with S9) from
urban soils, these results are biased by fact that most of
the urban data (>80%) was obtained from Japanese
assessments of MetOH extracts from contaminated
urban soils.
5.3. Relationships between Salmonella mutagenicity
and soil contamination
Several studies that employed the Salmonella
mutagenicity assay also examined levels of selected
contaminants previously shown to possess genotoxic
activity. Studies that examined soils collected from
industrial or urban areas frequently measured levels of
homocyclic PAHs, the products of high-temperature
combustion, including several known mutagens and
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 249
Fig. 4. Empirical relationship between the S9-activated Salmonella
TA98 mutagenic potency of soil extracts and soil PAH contamina-
tion (ppm dry weight). All values were log transformed to meet the
assumptions of least-squares regression. The overlaid information
shows the results of the linear regression analyses. All values are
from published soil mutagenicity assessments (see Appendix A).
Four observations represent mutation induction in Pseudomonas
putida by extracts of coal tar amended soil [90] (see Table 11). The
upper panel includes all available data. The lower panel does not
include the results of methanol extracts. Minimum, maximum, and
mean PAH contamination values are summarized in Table 4.
animal carcinogens (e.g., benzo[a]pyrene, dibenz[a-
h]anthracene). Sources of these PAHs include
industrial emissions (e.g., steel founding, coking),
mobile source emissions (e.g., automobile), and
natural emissions (e.g., forest fires, volcanoes). Three
studies measured levels of dinitropyrenes, a group of
compounds that include some of the most potent
bacterial mutagens ever examined, which are emitted
from heavy- and light-duty diesel vehicles including
automobiles, trucks and rail locomotives.
Empirical analyses showed a weak but significant
relationship between TA98 potency with S9 activation
and soil PAH concentration (r2 = 0.17, p < 0.0001).
Since PAHs are hydrophobic compounds that are
usually extracted from dry soil samples using a non-
polar solvent or solvent mixture such as hexane or
hexane/acetone [168–170], it is not surprising that the
strength of this relationship was significantly
improved when MetOH extract values were excluded
(r2 = 0.25, p < 0.0001). Despite the statistical sig-
nificance of these relationships, illustrated in Fig. 4, it
appears that the measured PAHs can only account for
17–25% of the variability in the S9-activated frame-
shift activity of soil extracts. Additional multiple
regression analyses investigated the relationship
between S9-activated TA98 mutagenic potency and
the concentrations of individual PAHs. Despite a high
degree of multicollinearity in the PAH data (e.g.,
average Pearson r = 0.91) that can complicate multi-
ple regression analysis [126], stepwise regression
analyses (e.g., Mallow’s Cp or MaxR selection. [127])
revealed a number of significant models. Using all
TA98 data the highest R2 was associated with a
model of mutagenic potency against soil concentra-
tions of fluoranthene and benzo[a]pyrene (R2 = 0.23,
F-ratio = 8.46, p < 0.0007). When MetOH extract
values were excluded, the results revealed a strong
empirical relationship between TA98 mutagenic
potency and soil concentrations of pyrene, benz[a]an-
thracene, and benzo[k]fluoranthene (R2 = 0.52, F-
ratio = 14.6, p < 0.0001). Examination of the stan-
dardized (i.e., unit-independent) regression coeffi-
cients, also referred to as beta coefficients, from the
aforementioned models suggest that the concentration
of individual PAHs would tend to over predict
mutagenic potency values relative to predictions
based on the concentration of the PAH mixture (i.e.,
total PAH). This seems reasonable since competition
for microsomal enzymes is thought to contribute to a
decrease in the mutagenic potency of PAH mixtures
such that they are below what would be expected if the
total mixture mutagenicity was simply the sum of that
expected from the mixture components [87]. Although
it is tempting to draw additional conclusions from the
beta coefficients, the high degree of multicollinearity
suggests cautious interpretation [126].
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345250
Fig. 5. Empirical relationship between the direct-acting Salmonella
TA98 mutagenic potency of soil extracts and DNP (dinitropyerene)
contamination (ppb dry weight). The DNP contamination value
represents the sum of 1,3-, 1,6- and 1,8-dinitropyrene. All values
were log transformed to meet the assumptions of least-squares
regression. The overlaid information shows the results of the linear
regression analyses. All data were collected from Watanabe et al.
[50,51,124] and Goto et al. [53] (see Appendix A). Minimum,
maximum, and mean values for DNP contamination are summarized
in Table 4.
Similar analyses revealed a strong empirical
relationship between TA 98 direct-acting mutagenic
potency and the soil concentration of dinitropyrenes
(r2 = 0.87, p < 0.0001). In contrast to the relationship
between PAH concentration and S9-activated frame-
shift activity, this relationship (Fig. 5) reveals that the
direct-acting frameshift activity of urban soil extracts
is largely determined by dinitropyrene contamination.
However, it should be noted that all of the observations
included in this analysis were obtained from Japanese
publications and the relationship may not be generally
applicable. One might expect the strength of this
relationship to be dependant on traffic density, the
proportion of diesel vehicles, and emission control
strategies employed in the soil collection area.
5.4. Mutagenic activity detected using other
Salmonella strains
Several recent studies employed metabolically
enhanced strains of Salmonella (e.g., YG1021,
YG1024, YG1026, YG1029) to examine the muta-
genicity of soil extracts. These strains possess enhanced
or reduced levels of enzymes required to metabolize
specific classes of chemical mutagens such as aromatic
amines and nitroarenes. The studies summarized in
Table 9 employed a variety of metabolically enhanced
strains that can assist in the identification of putative
mutagens in complex environmental extracts. For
example, strains YG1021 and YG1026 are TA98 and
TA100 derived strains, respectively, which possess
elevated classical nitroreductase (Cnr) activity that
dramatically increases their ability to detect some
nitroarenes including 1-nitropyrene, 2,6-dinitroto-
luene, and 2,4,6-trinitrotoluene [171,172]. Strains
YG1024 an YG1029 are TA98 and TA100 derived
strains that possess elevated O-acetyltransferase (OAT)
activity that dramatically enhances their ability to
detect samples containing direct-acting dinitroarenes
such as 1,8-dinitropyrene and S9-activated aromatic
amines such as 2-aminofluorene and 2-aminoanthra-
cene [173]. In addition, several mutant strains of TA98
have been constructed that lack nitroreductase (i.e.,
TA98NR) or O-acetyltransferase activity (i.e., TA98/
1,8-DNP6). When used in conjunction with the parent
strains, these strains also permit the identification of
complex samples containing aromatic amines and/or
nitroarenes.
The results summarized in Table 9 indicate that
nitroarenes make a major contribution to the
mutagenic activity detected in extracts of urban soils
collected in Japan and Germany [116,124]. Wesp et al.
[116] and Watanabe et al. [124] used metabolically
enhanced strains of TA98 and TA100 to determine that
polar aromatics such as nitroarenes make a major
contribution to the mutagenicity of soils collected
from highly urbanized sites (e.g., Mainz-Finthen
motorway, Kinki and Kanto megalopolises) that
receive heavy automobile traffic. The results obtained
using TA989NR and TA98/1,8-DNP6 to investigate
the direct acting activity of extracts from non-
agricultural soils collected in and around Livermore,
California (population �73,000) suggest that these
soils are also contaminated by nitroarenes [58].
Moreover, the mean TA98 potency value for these
soils of 0.17 � 0.03 reverants/mg is significantly
greater than the geometric mean TA98 potency for all
rural sites examined (i.e., 0.057 � 0.002). This may
not be surprising since some researchers have cited
airborne anthropogenic emissions as a major source of
soil mutagens [116], and the city of Livermore is
located predominantly downwind of the San Fran-
cisco-Oakland area (population �1,068,000).
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Table 9
Soil mutagenicity results obtained using metabolically enhanced strains of Salmonella
Sample(s) examined Salmonella strains employed Results obtained Reference
MetOH extracts of soils from
urban locations in Japan
TA98 derived strains YG1021a,
YG1024b, TA98NRc, and TA98/1,8-DNP6d.
Marked reduction in direct-acting
mutagenicity on TA98/1,8-DNP6.
Moderate increase on YG1021.
Marked increase on YG1024
[124]
DCM extract of creosote
contaminated soil
TA98 derived YG1041e and TA98NR,
and TA100 derived YG1042f
Marked reduction in direct-acting
mutagenicity on TA98NR. Some samples
show elevated mutagenicity on YG1041
and YG1042 without S9
[44]
DCM extract of creosote
contaminated soil
TA98 derived YG1041e and TA98NR,
and TA100 derived YG1042f
Detailed fractionation indicated that
putative mutagens include azaarenes
[22]
Thirteen soils collected from a
variety of contaminated sites
umu test in Salmonella typhimurium NM2009g Enhanced direct-acting activity in
extracts of TNT contaminated soils
and coking plant soils
[174]
Soils exposed to traffic exhaust
(German Autobahnen)
for up to 26 weeks
TA98 derived strains YG1021, YG1024, TA98NR,
and TA100 derived strains YG1026h and YG1029i
Response pattern in Salmonella strains
to the polar aromatic fraction indicates
a major contribution by nitroarenes
[116]
Roadside soils collected at 13
locations in Kurume City (Japan)
TA98 derived strain YG1041, TA100 derived strains
YG1042, TA1535 derived strain YG7108j,
and TA102 derived strain YG3003k
No responses in repair deficient YG3003
and YG7108. Substantial increases on
YG1041 and YG1042 indicative of
nitroarenes and/or aromatic amines
[122]
ACN extract of non-agricultural soils TA98 derived strains TA98NR and TA98-1,8-DNP6,
and TA100 derived TA100NRl
Marked reduction in activity mutagenicity on
TA98NR, TA100NR, and TA98-1,8-DNP6
without activation
[58]
a TA98 (hisD3052 TA1538 + pKM101) with classical nitroreductase on plasmid pYG216 [456].b TA98 with O-acetyl transferase on plasmid pYG219 [457].c TA98 lacking classical nitroreductase activity [458].d TA98 lacking O-acetyl transferase activity [459].e TA98 with classical nitroreductase and O-acetyl transferase on plasmid pYG233 [460].f TA100 (hisG46 TA1535 + pKM101) with classical nitroreductase and O-acetyl transferase on plasmid pYG233 [460].g TA1535/pSK1002 with pNM12 (enhanced O-acetyl transferase activity) [461].h TA100 (hisG46 TA1535 + pKM101) with classical nitroreductase on plasmid pYG216 [456].i TA100 (hisG46 TA1535 + pKM101) with O-acetyl transferase on plasmid pYG219 [457].j TA1535 (hisG46) lacking O6-methylguanine DNA methyltransferases [462].k Repair deficient version of TA102 (hisG428) [463].l TA100 lacking classical nitroreductase activity [458].
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345252
Tsukatani et al. [122] also employed a variety of
specially designed TA98 and TA100 derivatives to
investigate the mutagenicity of MetOH extracts from
urban soils in Japan. Strains YG3003 and YG7108,
which have enhanced sensitivity to oxidative muta-
gens and alkylating agents, respectively, showed no
mutagenic responses. However, extracts of samples
collected on roadway medians or adjacent to
intersections showed enhanced responses, both with
and without S9, on the OAT and Cnr enhanced TA98
and TA100 derivatives YG1041 and YG1042. The 4-
to 90-fold increase on YG1041 relative to TA98, and
the 6- to 77-fold increase on YG1042 relative to
TA100, confirm the involvement of both nitroarenes
and aromatic amines in the observed mutagenic
responses.
Additional studies employing metabolically
enhanced strains of Salmonella include the analyses
of extracts of creosote-contaminated soils by Hughes
et al. [44] and Brooks et al. [22], as well as analyses of
extracts of explosive-contaminated soils by Ehrlich-
mann et al. [174]. All three studies indicate that the
putative mutagens in these samples include nitrogen
heterocyclics, aromatic amines, and nitroarenes.
5.5. Miscellaneous Salmonella mutagenicity data
Table 10 summarizes the results of 14 studies that
were not used in the data analyses described in Section
6 [38,44,60,62,147–149,151,153,154,175–178]. Thir-
teen of the listed studies did not contain the
information required to calculate mutagenic potency
and/or express potency in revertents per equivalent dry
weight of soil. Most of these studies expressed the
results as Salmonella revertents per unit of extractable
organic matter (EOM in mg or mg per plate). Although
this is perfectly reasonable for studies that are
employing bioassay-directed fractionation to isolate
and identify the putative mutagens, it does not permit
cross-study comparisons of soils from different areas.
Conversion from revertents per unit EOM to revertents
per weight of soil requires the yield of EOM per unit
weight of soil. Only a sub-set of published studies
actually include this value (e.g., residue mg/mg soil
[42,48,54,115,165,166,179]). Readers should be
aware that some researchers employing the Salmo-
nella assay for examinations of soil extracts use the
term specific activity to refer to mutagenic potency in
revertents/mg EOM, and weighted activity to refer to
mutagenic potency in revertents per unit weight of soil
(e.g., [42,48,56,64,114,115,117,118,165,166,179]).
Although potency values in revertents/mg dry
weight were not available in these studies, and could
not be calculated using the published information, the
results described are consistent with the aforemen-
tioned data analyses. Those studies that examined
extracts of heavily contaminated (e.g., Superfund) soil
detected potent mutagenic activity [38,44,147,148,
151,153,177]. In addition, strong activity on TA98
with S9 generally corresponds to extracts of soils
known to be contaminated with PAHs or waste
materials that are likely to contain PAHs [38,147,
151,153,177]. It is interesting to note that the study by
Meloni et al. of an area described as a highly
contaminated waste disposal site in the province of
Padua did not detect any mutagenic activity (TA100
and TA98 without S9) in acetone and cyclohexane
extracts [178]. Rather, they detected significant direct-
acting frameshift mutagenicity in the acetone extract
of the uncontaminated control soil.
One study contained TA98 and TA100 results that
were included in the data analyses. However, the study
also employed additional Salmonella strains that are
not commonly used for the assessment of soils and soil
extracts (i.e., TA97a, TA102, TA104) [60]. The results
obtained revealed a significant response on TA97a, a
frameshift strain that carries a +1 frameshift at a run of
cytosines (i.e., hisD6610) [33]. This strain, known to
be more effective for the detection of metals and
some quinones, is sometimes used to supplement
TA98 [134,180]. No significant responses were
obtained on strains TA102 and TA104, strains with
an AT-rich mutation target (i.e., hisG428) that are
effective at detecting cross-linking agents such as
mitomycin C [33].
6. Other prokaryote or molecular in vitro assays
used for soil genotoxicity assessment
A number of studies have employed less popular
prokaryotic assays for soil extract genotoxicity
assessment. These include the SOS Chromotest, the
rec� differential survival assay in Bacillus subtilus,
the Microscreen phage induction assay, and the
Mutatox1 assay. Table 11 summarizes published soil
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Table 10
Published Salmonella mutagenicity results not used for detailed data analysesa
Site(s) examined Strains employed Results obtained Reference
Soils amended with motor oil TA1535 and TA1537 on aqueous extracts Detectable frameshift mutagenicity (�S9) in all samples [154]
Soils (2) from coke production area
(600–800 ppm PAHs).
TA98 on DCM extracts Significant frameshift activity with S9 [153]
Soils (3) contaminated by engine oils or
pesticides (2–20 ppm PAHs)
TA98 and TA100 on DCM or
acetone extracts
Significant frameshift activity (�S9 and +S9).
Significant base-pair activity without S9
[147]
Creosote contaminated soil (Superfund) TA98 and TA100 on DCM extracts Extracts of untreated waste positive on TA100 without S9 [44]
Petroleum contaminated soils
(Tatarstan Republic, Russia)
Salmonella TA98 and TA100
mutagenicity on aqueous extracts
Positive responses (both strains) enhanced by S9 activation [150]
Soils contaminated with petroleum refinery
effluent (Tatarstan Republic, Russia)
Salmonella TA98 and TA100
mutagenicity on aqueous extracts
No significant positive response [150]
Soils (2) contaminated with petrochemical wastes Salmonella TA98 and TA100
mutagenicity on ethyl ether/MetOH extracts
No significant positive response [62]
Creosote (1) and petroleum contaminated (4) soils TA98 and TA100 on silica/alumina
fractionated DCM extracts
Significant frameshift activity from polycyclic
aromatic fractions (+S9) and polar fractions (�S9)
[38]
Soils from industrial (22) and
non-industrial (30) sites
Salmonella TA98 and TA100 mutagenicity
on DCM/acetone extracts
91% of the industrial sites, 33% of the non-industrial
sites yield positive response (predominantly TA98 + S9)
[148]
Surface impoundment contaminated with
wood-preserving wastes
Salmonella TA98 and TA100 on a crude
extract (hexane/acetone, and DCM)
and several fractions
Basic extract elicited significant positive responses
in TA98 and TA100 + S9. Alumina fraction A2
elicited a strong response on TA98 + S9
[151]
Soil (waveland fine sand) column (25 cm) treated
with a mutagenic (TA98 + S9) sludge extract
Salmonella TA98 mutagenicity on aqueous
leachates and acetone/hexane extracts
Acidic leachates showed mutagenic activity (+S9).
Weak activity (+S9) in organic extracts of
column soil (top 7 cm)
[464]
Composite soil from agricultural fields irrigated
with industrial and domestic wastewaters
(Aligarh City, India)
Salmonella TA97a, TA102 and TA104 on
MetOH, acetone, and ACN extracts
Significant positive responses for all extracts on
TA97a (with and without S9). Slight dose-related
increases (not significant) for TA102 and TA104
[60]
Soil samples from oil fields in Kuwait Salmonella TA98 and TA100 on
DCM extracts
No significant positive response despite detection
of aromatics including benzo[a]pyrene
[155]
Soils from two toxic waste
disposal sites in Pavia (Italy)
Salmonella TA98 and TA100 (without S9)
on cyclohexane and acetone extracts
Significant positive response only for acetone
extract of control (uncontaminated) soil
[178]
Wood preserving bottom sediment (EPA K001) Salmonella TA98 on DCM/MetOH extracts Significant positive response with S9
(30%, v/v in S9 mixture)
[177]
Soil amended with treated municipal wastewater TA98 on DCM extracts of solid and
aqueous portion of soil samples
Greater direct-acting activity (per unit volume
of extract) associated with solid fraction
[175]
a The majority of the authors did not supply the information required to express potency values in revertents/mg dry soil. Standard plate incorporation assay, unless otherwise noted.
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4Table 11
Miscellaneous soil mutagenicity results — bacterial and molecular in vitro assay systems
Site(s) examined Bioassay employed Results obtained Reference
Storm-water runoff impoundment
(SWRI) wastea
Differential survival assay in rec�Bacillus
subtilus on DCM extracts
Marginal genotoxicity with S9
activation (<2 times background)
[49,166]
Combined APIb separator sludge
and slop-oil emulsion solids
Differential survival assay in rec�Bacillus
subtilus on DCM extracts
Moderate genotoxicity in acid fraction
with S9 activation (<5 times background)
[49,166]
Wood-preserving bottom sediment
(surface impoundment)c
Differential survival assay in rec� Bacillus
subtilus on DCM extracts
Positive response on acid fraction with
S9 activation. Approximately 10-fold lower
survival in rec deficient strains
[49,146]
Soils (2) contaminated with
petrochemical wastes
Differential survival assay in rec�Bacillus
subtilus exposed to ethyl ether/MetOH extracts
Positive (direct-acting) response
for one extract
[62]
Composite soil from agricultural
fields irrigated with industrial and
domestic wastewaters (Aligarh City, India)
Differential survival in recA, lexA, polA
mutants of E. coli K-12 exposed to MetOH,
acetone, or ACN extracts
Marked differential survival for all
extracts (MetOH > ACN> acetone).
polA mutants show largest difference,
followed by lexA and recA
[60]
Soils treated with coal tar extract Rifampicin resistance in Pseudomonas
putida (direct contact)
Changes in soil PAH contamination
unrelated to changes in genotoxicity
[90]
Soils from hazardous waste sites SOS Chromotest (SOS response induction in
E. coli) on DCM and cyclohexane extracts
S9-activated mutagenicity (at least 2-fold
increase over control) in most samples
[46]
Soils from a former manufactured
gas plant (MGP) site
SOS Chromotest (SOS response induction in
E. coli) on aqueous leachates
No positive response with or without S9 [189]
Soils collected near a coking facility
(600–800 ppm PAHs)
SOS Chromotest on DCM extracts Positive response with S9 [153]
Soils contaminated by engine oils or
pesticides (2–20 ppm PAHs)
SOS Chromotest on DCM extracts Marginal positive response without S9 [147]
Soils spiked with petroleum products
(e.g., kerosene)
SOS-Chromotest Pad (colourimetric SOS
Chromotest for solid samples)
Positive response for crude petroleum
and used motor oil only
[465]
Composted gasworks soil (Czech Republic) SOS Chromotest on DCM extracts Genotoxic activity with S9 only [188]
Eight soils collected from a military
(antitank) training area
SOS Chromotest on aqueous elutriates (pH 4.5) Several positive responses with and
without S9. S9 increased response in
3 samples. Three samples elicited
maximum IF > 1.5
[193]
Thirteen soils collected from a variety
of contaminated sites
SOS Chromotest and umu test in Salmonella
typhimuriumd on aqueous extracts and
concentrates of aqueous extracts
Potent direct acting samples from
TNT contaminated sites, moderate
responses on concentrates of extracts
from soils collected at coking plants
[174]
Wood preserving bottom sediment waste
(surface impoundment)
E. coli prophage induction assay on
several organic fractions
Genotoxic activity with S9 in the acid
and base fractions only
[151]
PCB contaminated soil E. coli prophage induction assay on DCM extracts Genotoxic activity with and without S9
(�500 pfu/g soil with S9e, �1700
pfu/g soil without S9)
[41]
Wood preserving bottom
sediment (EPA K001)
E. coli prophage induction assay
on DCM/MetOH extracts
Samples highly toxic. Some indication
of strong positive response (erratic)
[177]
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 255
Soil
sof
unknow
nori
gin
Let
hal
gen
etic
effe
cts
on
bac
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and
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32
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ore
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e
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cts
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[20
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[20
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ils
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ary
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nan
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).
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and
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sed
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ct
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genotoxicity assessments that employed other prokar-
yotic or molecular in vitro systems.
The first three aforementioned tests all rely on the
SOS response to DNA damaging agents [181,182].
The SOS Chromotest employs a variant of Escherichia
coli (strain PQ37) in which the production of b-
galactosidase is under the express control of the SOS
response to DNA damage, and SOS induction is
monitored colourimetrically [109,110,183]. Test
results are expressed as SOS induction factor (IF),
the ratio of toxicity-corrected SOS induction in the
samples relative to the solvent control, with sample
potency usually expressed as the SOS inducing
potency (SOSIP), the initial slope of the concentration
response relationship. A similar test, that has also been
used for soil genotoxicity assessment, is the Salmo-
nella umu test [111]. The test employs a plasmid (i.e.,
pSK1002) introduced into S. typhimuriumTA1535 to
place the production of b-galactosidase under SOS
control. The DNA repair or differential survival assay
in Bacillus subtilus investigates the differential
survival of SOS response deficient cells (e.g.,
rec�) in comparison to wild-type, repair proficient
cells [108,184,185]. The results are generally
expressed as a ratio of fractional survival between
the deficient and proficient cells after a selected
incubation period (e.g., 24 h at 37 8C). A variation on
this assay, recently employed for analysis of extracts
of agricultural soils, involved the use of various repair-
deficient mutants of E. coli strain K-12 [60]. The
Microscreen phage induction test assesses SOS
response induction by quantifying the frequency of
l prophage induction in E. coli WP2S [112,186]. The
results are generally expressed as plaque forming units
at a given concentration following overnight incuba-
tion of the WP2S reaction mixtures (i.e., test agent-
WP2S mixtures) with wild-type E. coli.
Each of these assays has been shown to respond to a
variety of base-pair and frame-shift mutagens, cross-
linking agents, intercalating agents, and DNA synth-
esis inhibitors [112,183–187], and they have been
successfully used to assess the genotoxic potential of
soil extracts. Eight published studies used the SOS
Chromotest to examine the genotoxic activity of soils
or soil extracts. Several studies noted positive S9-
activated genotoxicity in organic extracts (e.g., DCM
or cyclohexane) of soils contaminated by nearby
hazardous waste disposal facilities (i.e., Superfund
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345256
site), gasworks, or coking operations, [46,153,188].
These soils might be expected to contain S9-activated
PAHs [189], and analyses conducted in two of the
studies showed approximately 100–500 ppm PAHs at
the Superfund site and 600–800 ppm PAHs near the
coking operation [46,153]. A similar study that
investigated the genotoxicity of aqueous leachates
of soils collected near a former manufactured gas plant
(MGP) did not detect any genotoxic activity [189].
Although MGP are known to produce coal tars via the
pyrolytic destruction of coal [190,191], the negative
result is not surprising in light of the low water
solubility and high Kow of mutagenic PAHs [192]. Two
studies examined the genotoxic activity of soils
contaminated with a variety of petroleum products
(e.g., engine oils, kerosene). Malachova et al. detected
only marginal direct-acting genotoxicity in DCM
extracts of soils contaminated by engine oils or
pesticides (e.g., lenasil, trifluralin) [147]. Rojiekova et
al. used the SOS Chromotest solid phase test (i.e., SOS
Chromotest pad) to analyze soils amended with a
variety of petroleum products including kerosene,
used motor oil, and crude petroleum. The results
revealed direct-acting activity only for the crude
petroleum amended soils.
Robidoux et al. investigated the genotoxicity of
aqueous acid leachates from soils collected from an
antitank firing range [193]. The aqueous leachates,
suspected of being contaminated with polynitro-
organic (PNO) compounds associated with munitions
production and detonation, elicited positive direct-
acting responses. Most positive responses were
obtained in the absence of S9, and S9 addition
enhanced the responses to only three of eight leachates.
This is not surprising since a variety of PNO compounds
employed in munitions (e.g., TNT or 2,4,6-trinitroto-
luene), and their metabolites (e.g., 2-amino-4,6-
dinitrotoluene, 2,4-diamino-6-nitrotoluene, etc.), are
known to be direct-acting bacterial genotoxins
[159,194]. The highest response was obtained for a
soil shown to contain tetryl (N-methyl-N-2,4,6-
tetranitroaniline), an explosive compound that is
known to be a direct-acting Salmonella mutagen
(TA98 and TA100) [194]. Ehrlichmann et al.
employed the SOS Chromotest and the Salmonella
umu test to examine aqueous extracts of a variety of
soils contaminated by munitions and abandoned
armaments, coking plant wastes, and wood impreg-
nation wastes [174]. Munitions contaminated sites
known to contain TNT and other PNOs elicited potent
direct-acting responses on the umu and SOS assays,
and the response was increased when tested in an O-
acetyltranferase enhanced version of the umu assay in
Salmonella NM2009. Similar to the aforementioned
results obtained for aqueous extracts of soils from an
MGP site [189], aqueous extracts of soils collected
near coal mining or coking operations were not
genotoxic. However, the aqueous extracts from these
soils did elicit a moderate response in the umu assay
after 30-fold concentration on an ethylstyrene resin
[174].
Six studies employed a differential survival assay in
repair-deficient bacteria (e.g., the rec� assay in B.
subtilus) or the Microscreen phage induction assay in E.
coli WP2S (l) to investigate the genotoxicity of soil
extracts. Several studies used the rec� differential
survival assay in B. subtilus to investigate the genotoxic
activity of organic extracts (e.g., DCM, ethyl ether/
MetOH, acetone, ACN) from petroleum refinery storm-
water runoff impoundment waste (SWRI), combined
petroleum separator sludge and slop-oil emulsion
solids, wood preserving surface impoundment sludge,
soils contaminated with petrochemical wastes (Argen-
tina), and agricultural soil irrigated with industrial and
municipal wastewaters [49,60,62,146,166]. Analysis of
DCM extracts from the SWRI waste, combined
petroleum separator sludge and slop-oil emulsion
solids, and wood preserving surface impoundment
sludge showed marginal (<2-fold control), moderate
(<5-fold control), and potent (�10-fold control) S9-
activated genotoxic activity, respectively. The acid
fractions of both the combined petroleum separator
sludge and slop-oil emulsion solids and wood preser-
ving surface impoundment sludge elicited the strongest
responses [49,146,166]. One extract of the petrochem-
ical contaminated samples from Argentina yielded a
positive direct-acting response [62]. Aleem and Malik
[60] employed a host of repair deficient strains of E. coli
K-12 (i.e., polA, lexA, recA) to assess the genotoxic
activity of extracts from soils irrigated with industrial
and municipal wastewaters. The results showed marked
differential responses to MetOH, acetone, and ACN
extracts. MetOH extracts yielded the strongest
responses followed by ACN and acetone. polA mutants
were found to be more sensitive, followed by the lexA
and recA mutants.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 257
Three studies employed the E. coli Microscreen
prophage induction assay to investigate the mutageni-
city of organic extracts from soils contaminated with
chlorinated contaminants. Although the Salmonella
mutagenicity test is the most common test for the
examination of complex environmental samples,
several researchers have noted the increased sensitiv-
ity of the prophage induction assay for activity
associated with chlorinated compounds [41,195,196].
Two studies examined organic extracts of bottom
sediment from an impoundment at a wood preserva-
tion site [151,177]. The study by Cizmas et al. noted
that extracts of the wood preserving waste (WPW),
contaminated with numerous mutagenic PAHs (e.g.,
benz[a]anthracene, dibenzo[a,h]anthracene) and PCP,
induced a potent S9-activated response [151]. Sub-
sequent fractionation demonstrated that the acid and
base fractions were the most genotoxic. The study by
Hong et al. detected a strong S9-activated response in
an extract from a WPW-contaminated site. However,
the extracts were highly toxic and the responses were
erratic [177]. The third study examined DCM extracts
of PCB contaminated soils collected at a US Navy site
in Guam [41]. The results revealed that the soil
extracts elicited positive responses both with and
without S9, and the response was not reduced by
remediation.
The Mutatox1 assay, commercially available in kit
form from Azur Environmental, employs reversion of
a dark mutant (M169) of the luminescent bacteria
Vibrio fisheri to detect mutagenicity. Reversion of dark
mutants to luminescent wild type is detected using a
luminometer and the intensity of the luminescent
signal at a given concentration is directly related to the
reversion frequency and the mutagenic activity of the
tested sample [197,198]. The test, shown to respond to
a variety of base-pair and frame-shift mutagens, cross-
linking agents, intercalating agents, and DNA synth-
esis inhibitors [198], was employed in four studies that
examined the mutagenicity of soil extracts or aqueous
leachates [199–202]. Cook et al. and Picado et al.
examined aqueous extracts of soils from a military site
contaminated with petrochemicals, and soils from a
coke oven site in Portugal, respectively [199,201]. The
Cook et al. study detected a positive S9-activated
response for extracts of three soils contaminated with
middle distillate (e.g., diesel oil). Despite PAH
contamination above 1000 ppm, the latter study only
yielded an equivocal response with and without S9. As
mentioned earlier, the lack of response is likely due to
the inability of the aqueous solvent to extract and
concentrate the mutagenic PAHs in the tested samples
(e.g., benz[a]anthracene, benzo[a]pyrene, etc.). The
remaining two studies employed the Mutatox1
system to analyze extracts of soils contaminated with
munitions and related compounds [200,202]. The
study by Jarvis et al. noted that ACN extracts of soils
from a site used for ordnance destruction elicited
positive responses without S9, and these responses
were increased following composting treatment [200].
Examination of aqueous extracts from soils collected
at a former TNT production facility in Germany
showed that three soils elicited positive responses
without S9 [202]. Additional analyses revealed high
concentrations of both TNT (>100 ppm) and two
dinitrotoluenes (>35 ppm) in two of the three positive
aqueous leachates.
Three additional studies employed other prokar-
yote or molecular in vitro assays to examine the
genotoxic activity of soils or soil extracts. Alexander
et al. employed a forward mutation assay for
rifampicin resistance in Pseudomonas putida to
examine soil mutagenicity [90]. The assay used a
direct-contact approach to examine the induction of
rifampicin resistance following a 16- to 18-h incuba-
tion with a sterile soil treated with coal tar extract. The
results showed an initial increase in genotoxic activity
during bioremediation. Pererva et al. examined the
ability of soils to induce lethal mutations in two
bacteriophage (i.e., lambda and MS32) [203]. The
results showed that MS32 is more sensitive to the
effects of the soils examined. Shaw et al. employed an
in vitro reaction system containing calf thymus DNA
and an S9 activation mixture from 3-methylcholan-
threne induced rats to examine the ability of extracts
from coal-tar contaminated soils to induce the
formation of bulky DNA adducts [204]. The results
showed that adduct frequency is related to the
hydrocarbon concentration in the reaction mixture.
Collectively, the studies summarized in Table 11
demonstrate that several other bacterial assays can be
employed to assess the genotoxic hazards of soils. The
studies employed a range of assays to analyse aqueous
and organic extracts of soils contaminated with a
range of compounds such as PAHs, munitions and
explosive residues, petroleum distillates, and wood
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345258
preservatives. Although some of the assays are reliable
and extensively validated [183,187,205–207], and
have some advantages over the more popular plate
incorporation version of the Salmonella mutagenicity
test, there are also some noteworthy disadvantages.
For example, the SOS Chromotest offers the
convenience of miniaturization and the test can be
performed in only 2–3 h [208]. In addition, sample
sterility and the survival of the tester strain are not
required [109,110]. However, these advantages are
offset by a lack of strains for diagnosis of mutational
mechanism and compound metabolism, noted sensi-
tivity to toxic (i.e., bacteriostatic) effects, and
difficulties in handling coloured samples [208,209].
Moreover, some researchers have highlighted the fact
that divalent metals in aqueous extracts (e.g., Ni2+,
Zn2+, Cd2+, Cu2+, Pb2+, and Hg2+) may seriously
disrupt the performance of the enzyme assays that
form the cornerstone of the assay [208,210,211].
With respect to the Mutatox1 assay, the system is
hampered by a lack of mechanistic understanding
of the event(s) required to revert dark mutants
of V. fisheri [212]. The manufacturer claims that the
dark mutation is in the luminescence ‘‘regulatory
system’’ rather than one of the lux genes (i.e., luxA,
luxB) that code for the luminescent substance (i.e.,
luciferase) and its subunits [197]. The lux regulatory
system has been shown to be quite complex and
it is not clear whether induction of luminescence in the
V. fisheri dark mutant used in the Mutatox1 test
actually requires a mutational event. Ambiguity
surrounding the significance of a positive Mutatox1
result has led some researchers to recommend
confirmation with the Salmonella plate incorporation
assay [213].
7. Plant assays used for soil genotoxicity
assessment
Plant assays for the detection of mutagenic and
clastogenic effects have been in existence for many
years. For example, representatives of Allium (onion),
Tradescantia (Spiderwort), Crepis (Hawksbeard), and
Vicia were used as far back as the 1930s to assess the
clastogenic effect of ionizing radiation [214–218]. In
the 1960s and 1970s, representatives of several plant
genera such as Vicia, Allium, Hordeum (barley), Zea
(maize), Tradescantia, and Arabidopsis were adopted
for routine use in the detection of chemical mutagens
[219–226]. Several of the selected plants are readily
amenable to mutagenicity research (e.g., easy to
handle, sensitive, large chromosomes), and a small
number of assays have been validated and standar-
dized to stimulate routine use in the detection of
environmental mutagens [227–237]. These include the
Vica faba MN, chromosome aberration, and sister
chromatid exchange assays [229,234], the anaphase
aberration assay in Allium cepa root tips [231], the
Tradescantia MN and stamen hair mutation assays
[228,233,238], the gene mutation assay in Arabidopsis
thaliana [230,236], and the specific locus mutation
assays in Zea mays [232].
Several assays, such as the Tradescantia stamen
hair mutation assay, the Tradescantia MN assay, and
the Allium anaphase aberration assay have been
effectively employed to monitor environmental
mutagens in aqueous media (e.g., surface water,
effluent) as well as contaminated air [239–247]. Since
soil is the growth medium for most plants, it seems
logical that these assays should be amenable to the
assessment of soil genotoxicity.
Forty publications contained soil mutagenicity or
clastogenicity assessments based on a variety of plant
assays. With few exceptions, these publications
employed one of five plant assays: the Tradescantia
MN test, the Tradescantia stamen hair mutation test,
the anaphase aberration assay in Allium cepa, the
Arabidopsis gene mutation assay, and the waxy
locus mutation assay is Zea mays. Detailed descrip-
tions of these assays can be found in Ma et al.
[91,228,229,238,245], Gichner et al. [236], Grant et al.
[231,248,249], Kanaya et al. [234], Sandhu et al.
[235,250], Van’t Hof and Schairer [227], Underbrink
et al. [223], and Plewa [72,251].
In total, 462 observations were collected from 29 of
the 40 publications that contained sufficient data to
investigate patterns in the results for a given endpoint.
The most common endpoint encountered in the
literature, MN induction in Tradescantia, accounted
for 27.7% of the collected data. This was followed by
stamen hair mutation induction in Tradescantia,
which accounted for 24.4% of the data, and induction
of CAs in Allium root tips, which accounted for a
further 22.5%. Over half of the remaining 117
observations (�14% of the total) are Zea mays waxy
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 259
locus mutation data (forward and reverse), with the
remaining 52 observations including data from the
following assays: Arabidopsis gametic mutation
assay, Allium MN assay, Vicia MN assay, and the
Vicia sister chromatid exchange assay.
Soil mutagenicity or clastogenicity assessment
employing the aforementioned assays was conducted
using a variety of exposure methods including direct
soil exposure, soil slurry exposures, and exposures to
aqueous extracts/leachates or organic extracts (e.g.,
EtOH, DCM, or dimethyl sulfoxide). The majority of
the collected data (62.1%) were generated from
exposures to unaltered soils transported to the
laboratory or in situ plantings. Twenty-five percent
of the data represent exposures to aqueous extracts or
leachates, with a further 10.6% of the data generated
from dimethyl sulfoxide (DMSO) extracts. The
remaining 2.4% (11 observations) were generated
from EtOH or DCM soil extracts.
Since much of the published plant mutagenicity or
clastogenicity assessments only provide a single value
for each individual soil sample or soil extract/leachate,
it was not possible to calculate potency values from
concentration–response relationships. For the purposes
of this review, only maximum responses and the
responses to the negative control (e.g., tap water,
nutrient solution, uncontaminated soil) were recorded.
In addition, simultaneous observations of contamina-
tion by metals, PAHs and radionuclides were also
recorded. Appendix B contains the published plant
genotoxicity data discussed in the subsequent sections.
The collected data were divided into five categories
based on site descriptions: reference/control, agricul-
tural, industrial vicinity, heavily contaminated, and
geogenic. The first category represents reference sites
chosen for their distance from any known contamina-
tion source. Reference soils include greenhouse soils
[70,71], undescribed control plots [78], potting soil
[76], and garden soil [69]. Although these reference
samples were usually described as unimpacted by
vehicular and industrial emissions, such statements
were rarely verified. Alternatively, researchers work-
ing with aqueous leachates routinely employed a
reference solution (e.g., tap water, nutrient solution) as
a negative control [67,83,252]. Agricultural sites refer
to those used for cultivation or grazing. Examples
include agricultural soils from the Transcarpathian
region of the Ukraine [74], pest management research
plots [253], and pesticide contaminated farms [252].
This category includes sites that were treated with
selected pesticides in order to investigate their
genotoxic hazards. Sites labelled as industrial vicinity
are those that were chosen for their proximity to an
industrial setting; however, the site is not a known for
severe contamination due to direct industrial waste
disposal. These include sites near metal smelting
operations [70,76,82], as well as sites near petro-
chemical refineries and organic chemical production
facilities [68,71,83]. Sites specifically described as
hazardous waste dumping areas or areas amended with
hazardous materials received the designation heavily
contaminated [67,78,254]. Geogenic sites are those
that contain mutagenic substances thought to be of
geologic origin (e.g., the As-rich Carinthian sample of
Knassmuller et al. [76]).
Table 12 provides a breakdown, by site and assay,
of the plant mutagenicity and clastogenicity data
collected from the literature. The table indicates that
for some assays (e.g., Vicia faba SCE assay) the data
are very limited.
7.1. Allium cepa anaphase aberration data
The data summarized in Appendix B includes 104
observations of anaphase CAs in Allium cepa. Allium
species have large chromosomes (2n = 16) that are
well-suited to scoring of chromosome aberrations, and
tests for studying the genetic effect of chemicals on
Allium chromosomes date back as far as 1938
[249,255].
A detailed description of the suggested protocol
can be found in Grant [248,256]. Briefly, onion bulbs
or seeds are germinated and 1–2 cm long roots are then
exposed to soils, soil extracts, soil slurries, or soil
leachates for 2–24 h (usually one mitotic cycle). Roots
are then fixed, stained and 100 anaphase or telophase
cells scored for aberrations including chromosome
fragments and bridges. Some researchers also score
the frequency of vagrant chromosomes and multipolar
cells that are presumed to be the result of c-mitotic
events [257].
Almost 80% of these Allium aberration data
represent the results of direct soil exposures. The
remainder of the data represents the results of
exposures to aqueous or organic soil extracts.
Table 13 summarizes the Allium chromosome
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345260
Tab
le1
2
Bre
akdow
no
fth
ep
lant
assa
yd
ata
coll
ecte
dfr
om
the
lite
ratu
re
Sit
e/sa
mp
leca
teg
ory
Nu
mb
ero
fo
bse
rvat
ion
sfo
rea
chcy
togen
etic
/mu
tag
enes
isen
dp
oin
ta
Chro
mo
som
alab
erra
-
tio
ns
Gen
em
uta
tio
ns
Ind
uct
ion
of
mic
ron
ucl
eus
form
a-
tio
n
SC
Efo
rmat
ion
To
tal
All
ium
cepa
bA
rabid
posi
sT
rades
canti
ab
Zea
ma
ysb
Tra
des
canti
ab
Vic
iafa
ba
Vic
iafa
ba
Aq
cD
Cd
Org
eA
qD
CO
rgA
qD
CO
rgD
CA
qD
CO
rgA
qD
CA
q
Ref
eren
ce/c
on
tro
l2
31
23
11
26
92
16
0
Agri
cult
ura
l4
71
511
63
71
22
9157
Ind
ust
rial
vic
init
y9
48
91
15
06
16
28
17
81
75
81
96
Hea
vy
con
tam
inat
ion
13
23
46
16
84
3
Geo
gen
ic2
31
6
To
tal
16
81
78
39
12
84
17
65
54
47
27
17
78
46
2
aT
wo
ob
serv
atio
ns
of
mic
ron
ucl
eus
ind
uct
ion
inA
lliu
mce
pa,
and
four
obse
rvat
ions
of
chro
moso
mal
aber
rati
ons
induct
ion
inV
icia
fab
an
ot
incl
ud
ed[8
0,8
3].
bS
uffi
cien
td
ata
for
det
aile
dan
alyse
s.c
Aqueo
us
extr
act
or
leac
hat
e.d
Dir
ect
conta
ctex
posu
res
(i.e
.,no
extr
acti
on).
eO
rgan
icex
trac
t(e
.g.,
DM
SO
,E
tOH
,D
CM
).
aberration data for different sites categories and each
exposure type (i.e., extract or whole soil). The
geometric mean values for direct soil exposures
clearly indicate a trend showing increasing aberration
frequency with increasing contamination. This rela-
tionship is illustrated in Fig. 6 (upper panel). The
accompanying ANOVA results indicate a significant
empirical relationship (p < 0.0001) between max-
imum aberration frequency and site category. More-
over, the post-hoc mean comparisons revealed a
significant difference between each site category.
Although the geometric mean values in Table 13 show
some evidence of a similar relationship for the
aqueous/DMSO extract results, separate analyses
(not shown) revealed a borderline result with an F
ratio of less than 3.1 and an accompanying p-value of
0.053. Nevertheless, The lower panel of Fig. 6
indicates that the relationship between aberration
frequency and site category is maintained when all the
data are analyzed (p < 0.001). These results suggest
that the Allium root tip anaphase aberration test is well
suited to the detection of clastogens in soils, and to a
lesser extent soil extracts or leachates. It appears to
respond to a range of soil contaminants including
radionuclides [257], pesticides [75], and industrial
contaminants [67]. Moreover, the control values for
direct contact exposures are very stable (e.g.,
reference soil geometric mean = 1.65 � 0.11) and
the endpoint shows a 10-fold increase in response
across a range of soils. Kovalchuk et al. [257]
highlighted the sensitivity of the assays and noted that
the assay is more sensitive to ionizing radiation than
the Vicia faba MN test. Ma et al. [245] suggested that
the increased sensitivity of Allium might be due to the
greater total length of the diploid chromosomes and/or
the higher number of metacentric chromosomes. In
light of this sensitivity and the assays ability to
identify hazardous samples, it is somewhat surprising
that this rapid test has not been widely applied to
studies of environmental contamination.
Analyses of ANOVA outliers revealed eight
significant negative outliers from the relationship
between maximum aberration frequency and site
category for direct soil exposures (Fig. 6, upper panel).
All eight of the outliers are agricultural soils from the
Ukraine or Uzbekistan [74,75]. Unfortunately, the
authors of these studies do not provide detailed
descriptions of the relevant sites.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 261
Table 13
Descriptive summary of the collected Allium cepa chromosome aberration dataa
Category
Reference/control Agricultural Industrial vicinity Heavy contamination
Exposure to aqueous or DMSO extracts
N 2 9 9 3
Minimum 0.62 0.60 0.67 1.62
Maximum 0.81 2.98 6.33 3.20
Mean 0.72 1.70 3.11 2.34
S.E. 0.095 0.24 0.65 0.46
Geometric mean 0.71 1.54 2.53 2.25
Distribution NAb Yes Yes NA
Direct contact with contaminated soil
N 3 71 4 3
Minimum 1.59 0.60 3.10 13.70
Maximum 1.77 6.90 6.70 23.80
Mean 1.65 3.38 4.80 17.47
S.E. 0.058 0.19 0.91 3.19
Geometric mean 1.65 2.93 4.54 16.94
Distribution NA No NA NA
a Values are maximum aberrations per 100 cells for a given exposure/experiment.b Insufficient data for normality test.
Additional analyses of the results shown in the
lower panel of Fig. 6 revealed a total of ten outliers,
nine negative outliers and one positive outlier. Most of
the negative outliers are aqueous or DMSO extracts,
including extracts of heavily contaminated municipal
waste compost samples examined by Cabrera et al.
[67]. This pattern confirms that direct soil exposure is
a more effective means of assessing clastogenic
hazard. The remaining negative outliers are the
aforementioned Ukrainian and Uzbekistani agricul-
tural sites. It is interesting to note that the sole positive
outlier is a heavily contaminated soil from Chernobyl
that was found to contain over 6000 Bq/kg of 137Cs
[257]. The effect of 137Cs contamination on Allium
aberration frequency is illustrated in Fig. 7. The Figure
shows a striking relationship (r2 = 0.98, p < 0.0001)
between 137Cs contamination and the frequency of CA
in Allium exposed via direct soil contact for two
mitotic cycles.
7.2. Arabidopsis gene mutation data
Relatively few researchers have employed Arabi-
dopsis for studies of contaminated soils; however, one
interesting study used the Arabidopsis gene mutation
system to examine the effects of radionuclide-
contaminated soils [68,258]. Arabidopsis is a small,
diploid (2n = 10) species with a conveniently short life
cycle. Large numbers of plants can be grown in a small
area and a single plant can produce in excess of 50,000
seeds [230,236,259]. The test, originally developed by
Muller [260], and described in detail by Redei
[230,259], involves scoring lethal, chlorophyll defec-
tive embryos (i.e., seeds) in the siliquae (i.e., the dry,
elongated fruit) of plants grown from seeds exposed to
the test substance [236,249]. Embryonic mutations
one generation removed from the initial exposure are
usually scored as chlorophyll (e.g., white, yellow, or
pale green seeds) and/or structural aberrations (e.g.,
abnormally small and shrunken).
Kruikov et al. [258] used the Arabidopsis gametic
mutation system to examine the frequency of
dominant lethal mutations in embryos from plants
grown in soils contaminated with 137Cs. The results
showed an enhanced frequency of mutations in soils
with greater levels of 137Cs contamination. When
these data are combined with the spontaneous
Arabidopsis mutation frequency [68] and background
levels of 137Cs [257], the combined data (N = 4) shows
a strong empirical relationship between mutation
frequency and radionuclide contamination. Fig. 8
illustrates the relationship between the frequency of
siliquae segregating for dominant lethal mutations and
soil concentration of 137Cs. Although only four
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345262
Fig. 6. Box plots of published Allium cepa anaphase aberration data
for soils and soil extracts from heavily contaminated locations,
industrial sites, rural/agricultural sites, and remote sites. The over-
layed text shows the results of the ANOVA analysis for a site
category effect. Potency values were log transformed to meet the
assumptions of least squares ANOVA. The upper panel includes the
results of direct-contact assays only. The lower panel includes all
available published data (e.g., direct contact, aqueous extracts,
organic extracts). Refer to the Fig. 3 legend for a detailed description
of the box. Boxes labeled with different letters are significantly
different at p < 0.05 (Duncan multiple range test). All values and
data sources are available in Appendix B. The number of observa-
tions in each site category is available in Table 13.
Fig. 7. The empirical relationship between the frequency of ana-
phase aberrations in Allium exposed to radionuclide-contaminated
soils (direct contact assessments) and the level of 137Cs contamina-
tion (Bq/kg). All values were log transformed to meet the assump-
tions of least-squares regression. The overlaid information shows the
results of the linear regression analysis. All values were obtained
from Kovalchuk et al. [257] and are shown in Appendix B. Mini-
mum, maximum, and mean 137Cs contamination values are sum-
marized in Table 4.
Fig. 8. Empirical relationship between mutation frequency in Ara-
bidopsis thaliana exposed to radionuclide-contaminated soils (direct
contact) and the level of 137Cs soil contamination (Bq/kg). Back-
ground values for Arabidopsis mutation frequency and 137Cs con-
tamination are from Chroust et al. [68] and Kovalchuk et al. [257],
respectively. All other values were obtained from Kriukov et al.
[258]. All values were log transformed to meet the assumptions of
least-squares regression. The overlaid information shows the results
of the linear regression analysis. Minimum, maximum, and mean137Cs contamination values are summarized in Table 4. All values
are available in Appendix B.
observations were available, the figure illustrates an
extremely strong empirical relationship between
mutation frequency and 137Cs contamination. This
relationship, as well as the aforementioned relation-
ship for Allium, is not surprising since ionizing
radiation such as X-rays are known to induce
mutations at a variety of plant loci [73,225,261–264].
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 263
The study by Chroust et al. employed the
Arabidopsis mutation assay to examine the effects
of PAH contaminated soils collected from two
industrial areas [68]. The results revealed that plants
grown in PAH contaminated soils (0.2–1.1 ppm)
displayed a higher frequency of dominant lethals in
the collected embryos. However, analysis of the
published data failed to reveal a significant correlation
between mutation frequency and PAH contamination
(Spearman rank order r = 0.9, p = 0.083).
7.3. Zea mays waxy locus mutation data
The plant data summarized in Appendix B include
65 observations of single gene germinal mutations at
the waxy locus in haploid Zea mays microgameto-
phytes (pollen grains) following direct contact with
contaminated soils. The plant, also known as maize,
has been called the pillar of classical genetics and it
was extensively used in early studies of spontaneous
and induced mutations [251,265–267]. Maize is
diploid (2n = 20), has conveniently large chromo-
somes, its growth requirements are well-known, and
hundreds of varieties with defined genotypes are
available for research [251,268,269].
A complete description of the assay can be found in
Plewa [72,251]. Briefly, the reverse mutation test (i.e.,
mutant wx to Wx) employs iodine staining to reveal
revertent Wx pollen grains that stain blue-black, in
contrast to wx pollen that stain tan-brown [70–72]. In
the forward mutation test wx mutants are detected as
tan-brown in contrast to the blue–black Wx pollen
[253]. The assay can be conducted by treating kernels
or plants, and both acute and chronic treatments can be
Table 14
Descriptive summary of the collected Zea mays waxy locus mutation dat
Category
Reference/contro
Direct contact with contaminated soil
N 12
Minimum 1.74
Maximum 12.88
Mean 5.02
S.E. 0.89
Geometric mean 4.34
Distribution Yes
a All values are maximum mutation frequency (�10�5) for a given ex
employed [251]. The reviewed studies employed
chronic, direct contact exposures until anthesis (i.e.,
flowering), usually 12–14 weeks [70–72,270,271],
and most studies employed the W22 or M14 inbred
lines to score reverse mutation frequency at the wx-C
or wx-90 alleles [251].
Although mutagenicity assessment with the W22
line has been shown to be effective for the detection of
soil mutagens (e.g., [72,270,271]), the overall size of
the plant can place practical limits on the utility of the
assay for routine analyses. Moreover, although the
long maturation period of 12–14 weeks ensures a
lengthy chronic exposure, the interval between
experiment initiation and data collection can be
problematic. Consequently, Plewa and Wagner [272]
developed an alternative waxy locus mutation assay
employing Early-Early Synthetic corn, a plant that
matures in four weeks and is only 50 cm in height. The
system has been shown to be sensitive and responsive
to potent mutagens such as ethyl methanesulphonate
(EMS) and maleic hydrazide (MH), as well as soil
amended with sewage sludge [72,272]. One of the
reviewed studies employed the Early-Early system to
examine the mutagenic hazards of soils treated with
varying amounts of pesticides [84].
Despite the limited quantity of published data, the
collected values summarized in Table 14 shows an
interesting trend of increasing mean mutation fre-
quency with site category. Reference sites showed the
lowest mutation frequency, followed by agricultural
sites exposed to a variety of pesticides [253,270,271],
and finally contaminated industrial sites that received
contamination from petrochemical and metal refining
facilities [70,71]. The relationship between mutation
aa
l Agricultural Industrial vicinity
37 16
2.45 3.72
27.54 72.44
7.97 22.68
0.81 4.38
6.92 16.93
No Yes
posure/experiment.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345264
Fig. 9. Box plots of the published Zea mays waxy locus mutation
data for soils from industrial sites, rural/agricultural sites, and
remote sites (direct contact only). The overlayed text shows the
results of the ANOVA analysis for a site category effect. Potency
values were log transformed to meet the assumptions of least squares
ANOVA. Refer to the Fig. 3 legend for a detailed description of the
box. Boxes labeled with different letters are significantly different at
p < 0.05 (Duncan multiple range test). The data shown were
obtained from six published studies [70–72,84,270,271]. Most
assessments examined reverse mutation at the wx-C allele in homo-
zygous inbred strain W22. Exceptions are the sewage sludge
amended soils examined via reverse mutation at the wx-90 allele
in inbred M14 [72,272], and the pesticide treated soils examined via
forward mutation at the Wx allele in Early-Early Synthetic Zea mays
[84]. All values are available in Appendix B. The number of
observations in each site category is available in Table 14.
frequency and site category is illustrated in Fig. 9.
ANOVA results revealed a significant relationship
(p < 0.0001) between the maximum mutation fre-
quency and site category. Moreover, post-hoc analyses
of mean values revealed significant differences
(p < 0.05) between each of the site categories.
Analysis of the ANOVA residuals showed two positive
outliers, one agricultural site treated with 4.48 to
4.80 kg/ha of the herbicide mixture SD50093 (atra-
zine + cyanazine) [270], and one site 100 m from a oil
refinery complex in Illinois [71]. These sites yielded
waxy locus mutation frequencies of 27.8 and
72.5 � 10�5 respectively; values that are 5- to 15-
fold greater than their respective controls. It is not
immediately obvious why either of these locations
induced responses that were deemed to be significant
positive outliers, particularly in light of the fact that an
observation from the same industrial site in a previous
year showed a markedly low response that was
deemed to be a negative outlier [71,270]. A second
negative outlier corresponded to a control site from
Lower et al. [70] located in Granite City, Illinois
3.2 km from a lead smelter in Boss, Missouri.
It is interesting to emphasize that three studies
examining soils treated with a variety of herbicides,
insecticides, and fungicides frequently noted marked
increases in gametophyte mutation rate, despite the
fact that some of these pesticides were not found to be
mutagenic in either Salmonella or Saccharomyces
cerevisiae (reverse mutation at ade or trp loci).
Exceptions include the herbicides dicamba, metola-
chlor, alachlor, and procyazine, as well as the
insecticides fonofos, chlordane, heptachlor, and
terbufos [270,271], and the fungicide captan. The
latter compound, often used for seed treatment prior to
storage, is known to be mutagenic in a variety of assay
systems [195,273–275].
Rodrigues et al. [253] showed that heavy applica-
tions of pesticides for seed treatment and weed control
(e.g., cyanazine, metolachlor, diazinon, chlorpyrifos,
carboxin, and captan) resulted in a two-fold increase
in mutation frequency (�8 � 10�5) in comparison
to reference samples that did not receive any pesti-
cides (�4.5 � 10�5) and/or laboratory controls
(�3 � 10�5) grown in a commercial soil mixture.
Plewa et al. [270] showed substantial increases in
mutation frequency (i.e., 2- to 5-fold) in plants
exposed to soils where certain herbicides had been
applied. Application of the triazine herbicides such as
cyanazine or simazine, compounds that are not
mutagenic in Salmonella or Saccharomyces [276–
279], induced mutation frequencies of approximately
11–28 � 10�5 in comparison to control values of
approximately 3–5 � 10�5. Combinations of metola-
chlor and triazine herbicides also induced substantial
increases over the control. In addition, Gentile et al.
[271] noted significant (�2-fold) increases in muta-
tion frequency in plants exposed to soils where the
insecticides chlordane, ethoprop or heptachlor had
been applied. Although some of the pesticides studied
by Plewa et al. and Gentile et al. are clearly non-
mutagenic in Salmonella, it is interesting to note that
Plewa et al. [270] also demonstrated that Salmonella
incubation with a metabolic activation mixture
derived from Zea mays kernels results in strong
positive responses on TA100, TA98 and TA1537.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 265
7.4. Tradescantia stamen hair mutation data
Mutagenicity assay systems employing represen-
tatives of Tradescantia for chemical screening, as well
as in situ assessment of airborne mutagens, have been
available since the late 1960s and 1970s [223,225,
239,280–282]. The most popular assay systems, the
stamen hair mutation system and the MN assay in
meiotic pollen mother cells, are generally carried out
using sterile hybrid clones such as Tradescantia clone
4430, an interspecific hybrid of T. hirsutiflora and T.
subcaulis. The sterility of the clones is a convenient
feature that ensures genetic homogeneity in the
absence of mutagenic effects. A complete description
of both assays can be found in Ma et al. [91,228,
238,245,247], as well as important earlier works by
Underbrink et al. [223] and Sparrow et al. [225].
The collected plant data includes 113 observations
of stamen hair mutation frequency assessment in
Tradescantia. With three exceptions, all the reviewed
studies employed the aforementioned sterile hybrid
denoted clone 4430 [66,67,69–71,81,283]. One study
employed clone 02, also referred to as clone BNL02
(Brookhaven National Laboratory clone 02), a
putative diploid hybrid of T. occidentalis and T.
ohiensis [73], and two Russian studies employed an
unspecified isolate of T. poludosa [74,75]. The stamen
hair mutation assay or Trad-SHM is based on the fact
that stamen hair cells in clone 4430, and other clones
such as BNL02, are heterozygous for phenotypically
visible flower colour markers (i.e., blue-dominant and
pink-recessive) [91]. Briefly, cuttings of Tradescantia
clones heterozygous for the alleles controlling stamen
hair colour are exposed to aqueous extracts, organic
extracts diluted in an aqueous medium, whole soils, or
soil slurries for up to several days. Following a lag
period of up to 14 days (depending on treatment time)
stamen filaments are microscopically examined and
scored for pink mutations. The results are usually
expressed as mutation events per 1000 stamen hairs.
Although the most commonly employed protocols for
treatment in aqueous media use relatively short
exposure times between 6 and 30 h [66,67,69,81,
91,252,283], assessments of soils by direct contact,
and assessments of airborne mutagens often employ a
chronic exposure lasting for several days [227] or even
several weeks [70,71]. For example, in their studies of
sites in the vicinity of a lead smelter and a
petrochemical complex, Lower et al. conducted
lengthy 6–12-week direct contact soil exposures
[70,71].
A summary of the collected stamen hair mutation
data is provided in Table 15. Almost 75% of the
collected values were generated from direct soil
exposures. The remaining data includes exposures to
aqueous or organic (DMSO or EtOH) soil extracts
[67,284]. Variations in extraction and exposure
method, and background mutation frequency across
the various studies complicated comparisons across
site categories. For example, the direct soil exposure
data indicate that the mean mutation frequency in
plants exposed to agricultural soils is less than that for
the reference (i.e., control) soils (Table 15, middle).
However, closer examination of the agricultural
data (see Appendix B) indicates that the geometric
mean mutation frequency for the 11 Uzbekistani
and Ukrainian sites examined by Kurrinyi et al.
(1.7 � 10�3), although unusually low in comparison
with other published values, is significantly greater
than its matching reference values (i.e., 0.85 and
0.4 � 10�3) [74,75]. Both of the Kurrinyi et al. studies
employed an unspecified isolate of T. poludosa and it
seems clear that this isolate yields spontaneous and
induced mutation frequency values that are low in
comparison to those recorded by other researchers.
For example, the reference data from the studies
of Ichikawa and Ishii and Lower et al. [70,71]
provided mean mutation frequency values in the (1.8–
2.5 � 10�3 range, 2- to 6-fold higher than the Kurrinyi
et al. values. This required separation of the Kurinnyi
et al. [74,75] data from the other published data prior
to detailed data analysis.
The relationship between mean mutation frequency
and site category for the remaining direct contact data
is illustrated in Fig. 10 (upper panel). The figure and
accompanying ANOVA results indicate a significant
relationship (p < 0.0001) between mean mutation
frequency and site category (industrial and reference
only). Therefore, the direct-contact version of the
SHM assay with Tradescantia clones 4430 or BNL02
appears to be effective at discriminating between
background mutagenicity and levels associated with
sites near industries associated with the production
and emission of mutagenic substances (e.g., PAHs).
However, it should be noted that the data from
Ichikawa and Iishi [73], which were generated using
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345266
Table 15
Descriptive summary of the collected Tradescantia stamen hair mutation (SHM) dataa
Category
Reference/control Agricultural Industrial vicinity Heavy contamination
Exposure to aqueous or organic extract
N 2 6 17 4
Minimum 1.19 3.41 0.89 1.72
Maximum 1.29 7.20 5.82 2.36
Mean 1.24 4.68 2.80 1.94
S.E. 0.05 0.60 0.31 0.14
Geometric mean 1.24 4.50 2.50 1.92
Distribution NAb Yes Yes NA
Direct contact with contaminated soilc
N 23 11 50 No data
Minimum 0.40 0.27 1.65
Maximum 3.87 3.00 7.17
Mean 2.12 1.59 3.38
S.E. 0.17 0.22 0.18
Geometric mean 1.95 1.39 3.16
Distribution No No No
Direct contact: clones 4430 and BNL02 onlyd
N 22 No data 50 No data
Minimum 1.34 1.65
Maximum 3.87 7.17
Mean 2.20 3.38
S.E. 0.16 0.18
Geometric mean 2.09 3.16
Distribution No No
a All values are maximum mutation frequency (�10�3) for a given exposure/experiment.b Insufficient data for normality test.c Includes clone 4430, clone BNL02, and an unspecified isolate of T. poludosa.d Excludes values generated using an unspecified isolate of T. poludosa [74,75].
clone BNL02, do suggest that this clone may be less
sensitive than clone 4430. Clone BNL02 yielded lower
reference and industrial mean values and a subsequent
t-test comparing BNL02 and 4430 values showed a
significant difference (p < 0.02) between the mean
reference values, 1.78 � 10�3 � 0.07 and 2.49 �10�3 � 0.23 (direct contact only), respectively. In a
much earlier work comparing these two clones,
Sparrow et al. also noted that the mutation rate in
4430 following a chemical exposure was appreciably
higher than that of clone 02 [225].
The remaining 25% of the Tradescantia SHM data
(N = 29) are the results of assessments that employed
aqueous or organic soil extracts (Table 15). These
results, presented in Fig. 10 (lower panel), based on
data collected from five studies [66,67,69,81,283],
indicate that there is very little difference between the
mean mutation frequency across the different site
categories. Although the ANOVA analysis permitted
rejection of the null hypothesis at p < 0.004, the post-
hoc comparison of mean values indicates that the
results from heavily contaminated soils are not
significantly different from the results obtained for
reference soils. In addition, the mean of the rural/
agricultural soils is significantly greater than the mean
for the heavily contaminated soils (e.g., composted
municipal waste). This may be due to the fact that all
the agricultural/rural values reflect the mutagenicity of
DMSO extracts from soils irrigated with water from
the Queretaro River, a source that receives wastewater
inputs from a variety of industrial and domestic
sources. In addition, mine-dump sites with extremely
high levels of metals that have been shown to be
genotoxic in Tradescantia assays were categorized as
industrial [76,77]; however, the actual samples
examined contained low concentrations of metals
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 267
Fig. 10. Box plots of the published Tradescantia stamen hair muta-
tion data for soils and soil extracts from heavily contaminated sites,
industrial sites, rural/agricultural sites, and remote sites. The over-
layed text shows the results of the ANOVA analysis for a site category
effect. Potency values were log transformed to meet the assumptions
of least squares ANOVA. The upper panel includes the results of
assays with direct soil contact only. The bottom panel includes the
results of exposures to soil extracts or aqueous leachates only. Refer to
the Fig. 3 legend for a detailed description of the box. Boxes labeled
with different letters are significantly different at p < 0.05 (Duncan
multiple range test). The values presented were collected from 8
published studies [66,67,69–71,73,81,283]. All values, shown in
Appendix B, were generated using Tradescantia clones 4430 or
BNL02 [91,225]. Exclusion of clone BNL02 from the upper panel
yields a slightly stronger ANOVA result (r2 = 0.34, N = 45, F-
ratio = 22.3, p < 0.0001). Two studies that used an unspecified stock
of Tradescantia poludosa were not included in the figure or accom-
panying data analyses (see text and Table 15) [74,75]. The number of
observations in each site category is available in Table 15.
(e.g., Cr, Cd, Ni, Zn, Pb) and induced weak responses
[81]. Finally, variability in the Trad-SHM soil extract/
leachate data may also be due to variations in the
exposure duration employed in the reviewed studies.
Although continuous treatment for 30 h is generally
recommended for liquid samples [66,67,252,283], the
cited studies used a variety of exposure times between
6 and 30 h [69,81,252].
In general, the paucity of aqueous/organic extract
data, and difficulty in objectively classifying the study
sites complicated the interpretation of the data
presented in the lower panel of Fig. 10, and it is
not possible to draw any firm conclusions. However,
the small range in the maximum mutagenic response
to aqueous or DMSO extracts from a range of sites is
troubling. The assay does not appear to have a great
deal of dynamic range, and, moreover, there is a fair
degree of variation in spontaneous mutation frequency
across studies. The five studies discussed here show
negative control values that range from 1.9 to 3.49
mutations per 1000 stamen hairs [66,67,69,81,283],
and the positive controls maleic hydrazide and O-
phenylenediamine often yielded responses that are
less than 4-fold above the control.
7.5. Tradescantia micronucleus test data
The Tradescantia MN assay or Trad-MN, an assay
originally developed for the assessments of gaseous
and airborne mutagens [227,239,256], is often used
for investigations of contaminated aqueous media
such as surface waters or wastewaters [241,242,246,
247,285,286]. It is very popular, and its utility for
analyses of complex environmental samples has been
the subject of several review papers and international
evaluation exercises [78,228,235,238,246,250,287,
288]. The assay involves exposure of 15–30 cuttings
per treatment to the test material, and inflorescences
are subsequently fixed and early stage tetrads (i.e.,
meiotic products of spore mother cells) are stained
and scored for MN. Generally, 300 tetrads are scored
from each experimental group and the results are
expressed as MN per 100 tetrads [238]. Exposures
times vary widely depending on the nature of
the study and the media being examined. Typical
values employed for complex mixture analyses
include 3–12 h [69,83,238] for acute aqueous
exposures, 24–30 h for chronic aqueous exposures
[81,120], 3–24 h for in situ water monitoring [238],
up to 10 days for monitoring of contaminated air
[227], and 72 h for direct contact monitoring of
contaminated soils [76].
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345268
Table 16 summarizes the data from 17 published
soil genotoxicity assessments that employed the
Tradescantia MN test. Of the 128 observations
collected from the literature, approximately 63% used
the assay to assess the mutagenic activity of aqueous
soil leachates/extracts or DMSO extracts. These data
are almost equal amount aqueous extract/leachate
values and DMSO extract values. The remaining 37%
of the data was obtained from studies that used the
assay to examine the effects of direct soil exposures.
Analysis of the direct contact results, illustrated in
Fig. 11, show that although the null hypothesis of
equal mean values for the various sites categories was
rejected at p < 0.05, post-hoc comparisons of mean
values indicated that examinations of reference, rural/
agricultural, industrial vicinity, and heavily contami-
nated sites yielded similar levels of MN induction.
Even heavily contaminated Superfund samples such
as those investigated by Gill et al., which yielded clear
Table 16
Descriptive summary of the collected Tradescantia micronucleus inducti
C
R
co
Direct contact with contaminated soil
N 9
Minimum 2.
Maximum 8.
Mean 6.
S.E. 0.
Geometric mean 5.
Distribution Y
Exposure to aqueous extracts, DMSO extracts, or aqueous leachates
N 8
Minimum 1.
Maximum 6.
Mean 4.
S.E. 0.
Geometric mean 3.
Distribution Y
Exposure to aqueous extracts and leachates only
N 4
Minimum 1.
Maximum 4.
Mean 3.
S.E. 0.
Geometric mean 3.
Distribution N
a All values are maximum frequency of micronuclei in numbers per 1b Insufficient data for normality test.
positive responses (e.g., p < 0.05) for direct soil and
soil slurry exposures, only induced a maximum
response 3.1-fold above the control [78]. This site
is known to be contaminated with a variety of
hazardous materials including pesticides (e.g., hepta-
chlor, dieldrin) that have been shown to induce MN in
Tradescantia [289]. The study by Majer et al. of soils
contaminated with varying amounts of toxic and
genotoxic metals (e.g., As, Cd, Sb, Cr, Cu, Mn, Ni, Pb,
and Zn) showed numerous significant positive
responses (p < 0.05); however, even the most dra-
matic responses only reached 3-fold above the mean
control value (�5 MN per 100 tetrads) [77]. Thus,
despite the ability to discriminate between test
samples and reference samples within one study,
the dynamic range appears to be too narrow to show
cross-study differences between reference and test
soils. The one exception are the soils with geogenic As
contamination examined by Knasmuller et al. and
on (MN) dataa
ategory
eference/
ntrol
Agricultural Industrial
vicinity
Heavy
contamination
Geogenic
2 17 16 3
84 5.97 4.38 3.50 11.0
70 6.34 100.0 23.80 77.0
10 6.16 18.36 10.51 34.8
62 0.18 6.44 1.43 21.2
80 6.16 10.75 9.21 24.0
es NAb No Yes NA
21 36 14 2
50 2.70 1.06 2.50 3.07
57 15.19 15.30 43.0 5.57
07 7.07 5.95 8.28 4.32
52 0.65 0.52 2.70 1.25
78 6.52 5.18 6.28 4.14
es Yes Yes No NA
10 18 6 No data
50 4.40 1.90 2.50
87 9.64 15.30 43.00
28 7.46 6.74 11.56
74 0.54 0.78 6.32
77 7.26 6.01 11.77
A Yes Yes NA
00 tetrads for a given exposure/experiment.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 269
Fig. 12. Empirical relationship between the MN frequency in
Tradescantia exposed to aqueous soil leachates and the level of
PAH contamination (ppm dry weight). All values were log trans-
formed to meet the assumptions of least-squares regression. The
overlaid information shows the results of the linear regression
analysis. All values shown were obtained from four published
studies [63,83,120,475] (see Appendix B). The relationship is not
significant if the extreme value from Baud-Grasset et al. [120] is
removed. Minimum, maximum, and mean PAH contamination
values are summarized in Table 4.
Fig. 11. Box plots of the published Tradescantia micronucleus
induction data for soils from heavily contaminated sites, industrial
sites, rural/agricultural sites, remote sites, and sites contaminated
with geogenic metals (direct contact only). The overlayed text shows
the results of the ANOVA analysis for a site category effect. Potency
values were log transformed to meet the assumptions of least squares
ANOVA. Refer to the Fig. 3 legend for a detailed description
of the box. Boxes labeled with different letters are significantly
different at p < 0.05 (Duncan multiple range test). The values shown
were obtained from 15 published studies [63,66,67,69,76–
78,81,83,85,120,252,283,286,474]. All values are available in
Appendix B. The number of observations in each site category is
available in Table 16.
Majer et al. [76,77]. These soils (i.e., Feistritz,
Saualpe) contained natural levels of As between
150 and 1500 ppm, and additional experimentation
showed an 11-fold induction of MN could be elicited
by exposure to soils artificially spiked with 990 ppm
As (as As2O3) [77].
Separate analysis of the Tradescantia MN results
for aqueous or DMSO extracts only showed no
significant differences between the mean values across
the four site categories (not shown). Therefore,
although some studies have stated that the MN assay
in Tradescantia is a sensitive endpoint for the
detection of mutagenic activity in environmental
samples (e.g., water, effluents, air) [85,252,283,289],
the data collected and analysed in this review do not
support its utility for analysis of soil extracts and
aqueous leachates. This assertion is supported by the
Knasmuller et al. study that observed significant
positive MN responses for direct soil exposures, but
not for aqueous leachates [76]. The authors of that
study concluded that the Tradescantia MN assay is
‘‘less appropriate’’ for the detection of mutagenic
activity in aqueous soil extracts. However, it should be
noted that is not clear whether the lack of sensitivity
observed with aqueous soil extracts is related to the
exposure medium or the exposure duration. As already
noted, exposure times for examinations of aqueous
samples are usually 3–12 h [69,83,238], whereas the
exposure duration for the direct-contact assessments
described by Knasmuller et al. was 72 h.
The assertion that exposure duration contributes to
a lack of sensitivity in the Trad-MN assay on aqueous
soil extracts is supported by the results of Baud-
Grasset et al. [120]. The study, which employed a 30 h
exposure to an aqueous extract of a creosote
contaminated soil with over 5000 ppm PAH, showed
a large increase in MN frequency [120]. In addition,
empirical analysis of the Tradescantia MN data
showed a significant positive relationship between
MN induction frequency for aqueous extracts and
PAH contamination. However, it should be noted that
the relationship (r2 = 0.27, p < 0.04), illustrated in
Fig. 12, is heavily dependent on the single extreme
value from the Baud-Grasset study. If this point is
removed, the relationship is no longer significant
(r2 = 0.005, F = 0.07).
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345270
Data from the 33 samples examined by Knass-
muller et al. and Majer et al. were used to explore
empirical relationships between MN induction in the
direct-contact assay and soil metal contamination
(e.g., As, Zn, Pb, Cr, Cd, Ni, Sb, Ni) [76,77]. The
results, illustrated in Fig. 13, show significant
relationships between Tradescantia MN frequency
and the concentrations of eight metals: Cr, Pb, Cd. As,
Cu, Sb, Ni, and Zn. All relationships are significant at
p < 0.03 and r2 values range from a low of 0.16 for
cadmium and zinc, to a high of 0.42 for antimony. It is
interesting to note that the Majer et al. study from
which 18 observations were obtained, failed to detect a
significant relationship between MN induction and
metal concentration. An earlier study by Steinkellner
et al. confirmed that the metals As3+, Cd2+, Zn2+, and
Pb2+ all induce positive responses in the Tradescantia
MN assay. In addition, Knasmuller et al. [76] showed a
significant positive response to Ni2+ and Cr6+;
however, neither study was able to show a significant
positive response for Cu2+ or Sb3+.
The genotoxicity of many metals, including Sb and
Cr, is a complex and controversial topic. For example,
Cu2+ elicits a positive response in the Mutatox1 assay
and the SOS Chromotest [290], but a negative
response in the Salmonella mutagenicity test (TA98
and TA100) [290]. However, although this same study
failed to detect a significant Salmonella response for
Cd2+ and Zn2+, Pagano et al. detected a significant
positive response in Salmonella TA97 [180]. Cu2+ has
also been shown to induce chromosome damage in
some in vivo assays such as the chick embryo MN and
CA test [291]. The genotoxicity of antimony is also
controversial. Antimony (Sb3+) elicits negative results
in the Salmonella mutagenicity test, the tk mutation
assay in L5178Y cells, and the MN assay in mouse
peripheral blood (in vivo), but a positive clastogenic
response in human peripheral lymphocytes, and
positive responses in the rec� differential survival
assay, an SCE induction assay in V79 cells, and an in
vivo CA assay in mouse bone marrow (chronic 21-day
exposure only) [292–294].
7.6. Other plant assays used for soil mutagenicity
assessment
Table 17 includes assessments of soil mutagenic
activity that examined induction of MN, CAs and
SCEs in Vicia faba, and induction of MN in Allium
cepa. These endpoints are not as popular for soil
genotoxicity assessments as the aforementioned assays
in Tradescantia, Allium, Zea and Arabidopsis, and the
amount of published data was insufficient for detailed
analyses. Nevertheless, it is useful to include a
descriptive summary of the results obtained with these
less popular assay systems. Five studies, three of which
also used one of the more popular tests, employed the
MN assay in Vicia to examine contaminated soils
[68,76,79,82,83]. The study by Knasmuller et al.
examined MN induction in both Tradescantia and Vicia
following direct contact exposures to metal contami-
nated soils (72-h) [76]. Although the results revealed a
strong positive response for Tradescantia (up to 15-fold
above control), the soils did not induce a significant
positive response in the Vicia assay. Three other studies
of metal contaminated soils revealed significant
increases in MN or CA frequency in Vicia exposed
to aqueous extracts of waste heap soil collected adjacent
to a chromium processing facility (Cr�3 ppm), tannery
waste leachate (Cr �86 ppm), or flyash-amended soil
(direct-contact) examined before and after composting
(Cr �30 to 130 ppm) [68,80,82]. The latter study also
observed a significant increase in the frequency of
chromosomal and mitotic aberrations [79]. It is
interesting to note that although all of these studies
detected high concentrations of chromium, and the
authors of the flyash and tannery waste studies
highlighted chromium as the probable source of the
genotoxic activity [82], Knasmuller et al. could not
induce MN in Vicia using exposures to Cr(III)Cl3 or
Cr(VI)O3 [76]. The mutagenicity of chromium VI has
been well studied and it is known to induce significant
positive responses in numerous assay systems (e.g.,
Salmonella mutagenicity, prophage induction, murine
peripheral blood MN) [112,180,290,295,296].
In an examination of soils contaminated with
organic contaminants, Cotelle et al. noted that the
Vicia MN assay was more sensitive than MN induction
assays in Allium or Tradescantia for the detection of
mutagenicity in a leachate sample from a soil
contaminated with PCBs and solvents (e.g., toluene,
benzene, chloroform) [83]. However, analysis of a
leachate of a PAH contaminated soil revealed similar
sensitivity for the three endpoints. A similar analysis
of an aqueous leachate from an industrial soil
contaminated with PAHs revealed a significant
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 271
Fig. 13. Empirical relationships between published MN frequency in Tradescantia (direct contact exposures) and the level of soil contamination
with various metals (ppm dry weight). All metal concentrations are total extractable metal, and all values were obtained from two published
studies [76,77] (see Appendix B). All values were log transformed to meet the assumptions of least-squares regression. The overlaid information
shows the results of the linear regression analysis. Minimum, maximum, and mean metal contamination values are summarized in Table 4.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345272T
able
17
Mis
cell
aneo
us
soil
muta
gen
icit
yre
sult
s—
pla
nt
assa
ys
Sit
e(s)
exam
ined
Bio
assa
yem
plo
yed
Res
ult
so
bta
ined
Ref
eren
ce
So
ils
(29
)h
igh
lig
hte
dfo
rh
igh
con
cen
trat
ion
s
of
toxic
met
als
(US
A).
Her
itab
le(p
aren
t-o
ffsp
rin
g)
min
isat
elli
tem
uta
tio
ns
inTa
raxa
cum
offi
cin
ale
.
Sig
nifi
cant
posi
tive
corr
elat
ion
bet
wee
nm
uta
tion
rate
and
leaf
met
al(F
e,N
i,M
n,
and
Cr)
con
cen
trat
ion
[29
8]
So
ilam
end
edw
ith
mun
icip
alsl
ud
ge
(Fra
nce
)S
om
atic
mu
tati
on
s(a
þ 1=a
1aþ 2=a
2
syst
em)
inN
ico
tia
na
taba
cum
exp
ose
d
tosl
ud
ge
amen
ded
soil
s.D
irec
tco
nta
ctas
say
No
sig
nifi
can
tin
crea
sein
som
atic
mu
tati
on
rate
[89
]
So
ils
amen
ded
wit
hco
alfl
yas
h(p
ow
er
pla
nt
ince
ntr
alIl
lin
ois
)
Nucl
ear
alte
rati
ons
inm
aize
(mea
sure
d
via
flow
cyto
met
ry).
Dir
ect
conta
ctas
say
Incr
ease
dfr
equen
cyo
fnucl
ear
alte
rati
ons
asso
ciat
edw
ith
fly
ash
trea
ted
soil
s
[29
9]
Pes
tici
de
con
tam
inat
edag
ricu
ltu
ral
soil
s(U
SS
R–
Uzb
ekis
tan)
CA
sin
roo
tti
ps
of
Go
ssyp
ium
hir
sutu
m
(co
tto
n).
Dir
ect
con
tact
assa
y
Her
bic
ide
use
asso
ciat
edw
ith
ah
igh
freq
uen
cyo
f
aber
rant
cell
san
ddec
reas
edm
itoti
cin
dex
[29
7]
Met
alco
nta
min
ated
soil
sn
ear
stee
lwo
rks
(Ita
ly)
AF
LP
and
FC
Man
alyse
so
n
Tri
foli
um
rep
ens
(dir
ect
con
tact
)a
Sig
nifi
can
tn
ucl
ear
alte
rati
on
sfo
llow
ing
a1
5-d
ayex
po
sure
[30
0]
aA
mpli
fied
frag
men
tle
ng
thp
oly
mo
rph
ism
(AF
LP
)an
dfl
ow
cyto
met
ry(F
CM
).
increase in SCE frequency in Vicia, but no significant
increase in MN frequency [68].
Table 17 also summarizes studies that employed a
variety of other plants assays to investigate the
mutagenic hazards of contaminated soils in France
[89], Uzbekistan [297], the United States [298,299],
and Italy [300]. Chenon et al. employed a reverse
somatic mutation assay in Tobacco (Nicotiana
tabacum) that assesses mutations at two chloroplast
differentiation loci a1 and a2 [89,301]. The assay
system, initially developed by Dulieu [301], examines
reversions of greenish-yellow double heterozygotes
(aþ1 =a1 aþ2 =a2) to green, and the clonal expansion of
green cells reveals green spots on greenish-yellow
leaves [301]. The test can score a very large number of
cells on each individual leaf and has been shown to be
very sensitive to low levels of atmospheric pollutants
[89,302]. However, direct contact with municipal
sewage sludge amended soil for 45 days did not
significantly increase the somatic mutation rate over
background [89]. Abdullaev et al. employed a root tip
CA assay in Gossypium hirsutum (cotton) to
investigate the mutagenic effects of repeated herbicide
(i.e., cotoran also known as fluometuron and toluin
also known as trifluralin) applications [297]. Inspec-
tion of primary sprout rootlets from seeds of plants
raised in agricultural areas that received repeated
pesticide applications revealed an increase in aberra-
tions over background. In addition, the aberration
frequency increased with each successive year of
application. These results are not surprising since both
herbicides have been reported to induce chromosome
damage in plants [303,304].
Two studies used flow cytometry to investigate
nuclear alterations in Trifolium repens (white clover)
and Zea mays (maize) exposed to soils collected near a
steel founding operation, and soils amended with coal
fly ash, respectively [299,300]. The flow cytometer
was employed to measure the fluorescence intensity
and DNA content of nuclei stained with fluorochromes
such as DAPI (4,6-diamidino-2-phenylindole) or
propidium iodide [305]. The results revealed nuclear
alterations in maize following exposure to soils
amended with high levels of coal fly ash (70 t/ha)
[299]. The soil collected near a steel founding
operation was found to induce significant quantities
of debris (e.g., broken or disrupted nuclei) in clover
following direct contact for 15 days [300]. The latter
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 273
study also used AFLP (amplified fragment length
polymorphism) analysis to assess DNA sequence
changes [306,307] in clover. The results revealed a
significant effect from heavy metal contaminated soils
collected close to the steel founding operation [300].
A recent study by Rogstad et al. investigated the
mutation rate at minisatellite loci in dandelions
(Taraxucum officinale) exposed to soils contaminated
with a variety of toxic metals [298]. More specifically,
VNTR (variable number tandem repeat) DNA probes
were employed to assess parent-offspring transmis-
sion (i.e., leaf versus seed) of mutations in specimens
collected from 16 regions with various levels of metal
contamination (e.g., Cr, Cd, Pb, Mn, Zn, Ni, Cu, and
Fe). Although the authors did not detect any empirical
relationships between mutation rate and metal
concentration in the soil, they did detect significant
relationships between leaf metal concentration (i.e.,
Cd, Fe, Mn, Ni) and minisatellite mutation rate [298].
8. Other eukaryotic assays used for soil
genotoxicity assessment
8.1. In vitro eukaryotic assays
Table 18 provides a summary of 10 studies that
employed a variety of other eukaryotic in vitro assays to
assess the genotoxicity of contaminated soils or soil
extracts [40,42,49,54,65,116,146,166,308,309]. Seven
of these studies employed yeast or fungal (e.g.,
Aspergillus sp., Saccharomyces sp.) assays to assess
mutagenic and/or clastogenic effects. These simple
eukaryotic organisms are easy to culture and maintain,
and have been widely used in genetic research [310].
They have the distinct advantage of being able to exist
in both haploid and diploid state, and consequently can
provide a comprehensive range of endpoints including
forward mutation, reverse mutation, mitotic recombi-
nation, aneuploidy, and gene conversion [94,311–316].
Several works by Donnelly et al. and Brown et al. used
haploid Aspergillus nidulans for the detection of
forward mutations at methionine suppressor loci
[94,95,315], and/or diploid Aspergillus nidulans for
the detection of mitotic recombination and non-
disjunction events [314,316], to examine soils con-
taminated with wood preserving wastes [42,49,317],
refinery wastes [49,166], and munitions [54]. The
results obtained show that exposures to extracts of
agricultural soils can yield up to 3.5 methionine
suppressor mutants per 106 survivors. Although this is
significantly above the solvent control value of
approximately 0.7 mutants per 106 survivors (without
S9), it is substantially lower than that obtained
following exposures to extracts of soils amended with
a variety of hazardous wastes [40]. Examinations of
soils amended with refinery wastes or wood preserving
wastes revealed significant levels of mutagenic activity
in both the haploid and diploid assay. Extracts of waste-
amended soils yielded approximately 100 to 300
mutants (per 106 survivors) for refinery wastes (i.e.,
storm-water runoff impoundment waste, separator
sludge and slop-oil emulsion solids), and approxi-
mately 30 to 174 mutants (per 106 survivors) for wood
preserving wastes [42,49,166]. Additional Salmonella
mutagenicity analysis of the soils amended with wood-
preserving wastes suggested that the Aspergillus
mutation endpoint may be more sensitive [42].
Although the refinery wastes might be expected to
contain PAHs [318], refinery amended soils did not
show increased mutagenic activity in the presence of
S9. However, addition of S9 did increase the mutagenic
activity of the soils amended with wood preserving
wastes, and additional chemical analysis did identify
mutagenic PAHs and heterocyclics compounds (e.g.,
fluoranthene, dibenzothiophene) [42,319].
Examination of extracts from the soils amended with
refinery wastes also elicited moderate mutagenic
activity (<3-fold above control) in the Aspergillus
diploid assay (e.g., mitotic cross-overs, non-disjunc-
tion) [49]. Testing of extracts from soils amended with
wood-preserving wastes in the same diploid assay
system yielded responses 2- to 4-fold above the control
[49]. Additional examinations of the unaltered wood
preserving waste (i.e., a soil/sludge mix from a waste
lagoon) revealed extreme mutagenic activity in the
haploid assay with S9 activation (i.e., >20,000 induced
mutants per 106 survivors), and moderate clastogenic
activity in the diploid assay (i.e., induced segregation
index <5-fold above control) [49,317]. Comparison of
various waste fractions showed that the acid fraction
(without S9) induced the major chromosomal abnorm-
alities and the base fraction induced minor deletions or
insertions (without S9). In contrast, the mutagenic
endpoint in haploid Aspergillus was more sensitive to
the neutral fraction (with and without S9). However,
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
27
4
Table 18
Miscellaneous soil mutagenicity results — eukaryotic in vitro assays
Site(s) examined Bioassay employed Results obtained Reference
Combined APIa separator sludge and
slop-oil emulsion solids
Aspergillus nidulans haploid forward
mutation assayb on DCM extracts
Potent mutagenic activity with and without S9.
Potency �109–180 mutants (per 106 survivors)
per gram soil. Slight increase with S9
[49,166]
Aspergillus nidulans diploid mutation
assayc on DCM and MetOH extracts
Moderate genotoxic activity in several fractions
(<3 times background)
[49,166]
Storm-water runoff impoundment
(SWRI) wasted
Aspergillus nidulans haploid forward
mutation assay on DCM extracts
Potent mutagenic activity with and without S9.
Potency �127–308 mutants (per 106 survivors)
per gram soil. No increase with S9
[49,166]
Aspergillus nidulans diploid mutation
assay on DCM and MetOH extracts
Moderate genotoxic activity in several fractions
(<3 times background)
[49,166]
Agricultural soils
(row crops and grazing land)
Aspergillus nidulans haploid forward
mutation assay on DCM extracts
Weak activity with and without S9. Higher
activity on grazing land. Potency �0.04–3.5
mutants (per 106 survivors) per gram soil
[40]
Wood-preserving bottom sediment
(surface impoundment)e
Aspergillus nidulans haploid forward
mutation assay on DCM extracts
Potent mutagenic activity with and without S9.
Potency �27–56 � 103 mutants (per 106 survivors)
per gram soil. Significant increase with S9
[42,49,317]
Aspergillus nidulans diploid mutation
assay on DCM and MetOH extracts
Positive response on acid, base, and neutral
fractions. ISIf 2–5 times control
[42,49,317]
Soils amended with wood
preserving waste
Aspergillus nidulans haploid forward
mutation assay on DCM extracts
Potent mutagenic activity with and without S9.
Potency �30–174 mutants (per 106 survivors)
per gram soil. Significant increase with S9
[42,49]
Aspergillus nidulans diploid mutation
assay on DCM and MetOH extracts
Positive response on acid, base, and neutral
fractions. ISIf 2–4 times control
[49]
Soils from surface impoundments
contaminated with munitions wastewater
Aspergillus nidulans diploid genotoxicity
assay on DCM and MetOH extracts
Positive response for soil from two impoundments.
2- to 4-fold above background
[54]
Urban soils from Yana, Sesavtchy,
Lokorsko (Bulgaria)
Saccharomyces cerivisiae (diploid) D7
and D7ts1g mutation and mitotic
recombination assay on toluene extracts
Dose related increase in mutagenic and
recombinogenic activity in D7ts1 strain from the
least contaminated (Lokorsko) to the most contaminated (Yana)
[308]
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 275
So
ilfr
om
Mai
nz-
Fin
then
mo
torw
ay(G
erm
any
)
SC
E’s
incu
ltu
red
hu
man
lym
pho
cyte
s
expose
dto
var
ious
org
anic
frac
tions
Doubli
ng
of
SC
Efr
equen
cyb
ypola
rar
om
atic
frac
tion
[11
6]
Agri
cult
ura
lso
ils
(Ira
n)
CA
shan
dS
CE
siin
V7
9an
dC
HO
fib
rob
last
sex
po
sed
toM
etO
Hex
trac
t
Hig
hle
vel
so
fC
As
and
SC
Es
[30
9]
So
ils
con
tam
inat
edw
ith
mu
nit
ion
sw
aste
Mu
tati
on
sat
HP
RT
locu
sin
Chin
ese
Ham
ster
V79
cell
so
nE
tOH
:DM
SO
(1:1
)ex
trac
ts
Mu
tag
enic
ity
up
to1
0-f
old
above
con
tro
lfo
r
con
cen
trat
ion
sth
atd
idn
ot
red
uce
clo
nin
gef
fici
ency
[65
]
aA
mer
ican
Pet
rolu
emIn
stit
ute
.b
Fo
rwar
dm
uta
tio
nas
say
(met
hio
nin
eau
xo
tro
ph
s)th
atd
etec
tso
ne
of
sever
alsu
ppre
sso
rm
uta
tio
ns
[46
6].
cD
etec
tsm
itoti
ccr
oss
-over
s,non-d
isju
nct
ion,
colo
ny
colo
ur
muta
tions,
and
dom
inan
tle
thal
s.F
or
det
ails
,se
e[3
14
,31
6].
dS
lud
ge
coll
ecte
dfr
om
the
sid
eo
fse
ttli
ng
bas
inth
atre
ceiv
esp
etro
leu
mre
fin
ery
run
off
.e
Slu
dg
e/so
ilm
ixfr
om
the
per
iph
ery
of
aw
oo
dp
rese
rvat
ion
lag
oo
n.
fIn
du
ced
seg
reg
atio
nin
dex
.g
ts1
mu
tati
on
,h
om
ozy
go
us
ind
iplo
idD
7,
incr
ease
sth
ep
erm
eabil
ity
of
the
cell
san
dco
nfe
rsa
5–
7-f
old
incr
ease
inse
nsi
tiv
ity
[32
0].
hC
hro
moso
me
aber
rati
ons.
iS
iste
rch
rom
atid
exch
ang
es.
although this study detected differences in the response
patterns for Salmonella, B. subtilus (differential
survival), and Aspergillus, the authors’ described the
overall assessments of waste mutagenicity as ‘‘compar-
able’’ [317].
Donnelly et al. also used the diploid assay to
examine DCM and MetOH extracts of soils from
surface impoundments receiving munitions waste-
waters [54]. For two of the four impoundments
examined, the results revealed 2- to 4-fold increases in
clastogenic effects over that observed for an extract of
background soil. Chemical analysis revealed TNT
concentrations of 225–1290 ppm for the two sites that
yielded positive samples.
Terziyska et al. employed a permeable strain of
Saccharomyces cerivisiae to examine the mutagenic
and clastogenic activity of toluene extracts of several
soils collected from urban locations in Bulgaria [308].
The strain employed in the study, S. cerivisiae D7ts1,
displays increased permeability and substantially
enhanced sensitivity to a variety of chemical mutagens
over that observed for the standard D7 tester strain
[96,320]. The soil results confirmed the increased
sensitivity of the D7ts1 strain, and showed a dose-
related increase in mutagenic and recombinogenic
activity from the least contaminated soil, collected at
Lokorsko, to the most contaminated soil collected at
Yana. Comparison with the negative Salmonella
mutagenicity results suggested enhanced sensitivity
of the S. cerivisiae D7ts1 assay.
Only three published studies employed mammalian
cells to assess the mutagenicity of contaminated soils
and soil extracts [65,116,309]. Zia’ee and Sabouni
employed Chinese Hamster ovary (i.e., CHO) and V79
lung cells to investigate the genotoxicity of a MetOH
extract of an agricultural soil [309]. The results
revealed that low doses of the MetOH extracts induced
both CAs and SCEs. However, they did not detect a
significant induction of alkali-labile DNA damage.
Berthe-Corti et al. employed the hprt assay in V79
cells to investigate the mutagenicity of EtOH/DMSO
extracts of a soil contaminated with munitions
compounds including TNT and hexogen (hexahy-
dro-1,3,5-trinitro-1,3,5-triazine) [65]. The results
revealed activity as high as 10-fold above the control
for concentrations that did not reduce cloning
efficiency. One study employed an SCE assay in
cultured human lymphocytes to examine the geno-
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
27
6
Table 19
Miscellaneous soil mutagenicity results — eukaryote in vivo assays
Site(s) examined Bioassay employed Results obtaine Reference
Soil from Mainz-Finthen motorway (Germany) Bone marrow MN in mice exposed to
organic fractions (oral gavage)
No increase in NPCE (micronucleated
polychromatic e ythrocytes) observed
[116]
Coal tar amended soil, fresh or aged for 9 months Bulky DNA adducts in liver and lung
of F344 rats exposed via diet for 17 days
Highly significa t increase in adduct
frequency in lu and liver. Lung
3-fold greater t n liver. Aging has no effect
[326]
Soils from urban/industrial sites (Czech Republic) Somatic mutation and recombination test
in Drosophila melanogastera on aqueous
and DCM extracts
Significant posi e response for
four of nine DC extracts and six
of eight aqueou extracts
[68]
Soils from swampy area near concrete
factory (Kazakhstan)
Somatic mutations in Drosophila
melanogaster exposed to organic extracts
No significant e ect [321]
Soils of unknown origin Somatic mutations in Drosophila melanogaster
(wing spots) exposed to various soils (12)
No significant e ect [203]
Radioactive soils (7000 Bq/kg) from Chernobyl Dominant lethals and sex-linked recessive
lethals in Drosophila melanogaster
14-day exposur contributed to an
increase early e bryonic dominant
lethals and sex- nked recessive lethals
[324]
Soil amended with municipal sludge (France) MN in Xenopus laevis exposed to
aqueous leachates
Significant effe at several
elevated leacha concentrations
[89]
Two soils contaminated with metals
and solvents, or PAHs (France)
MN in Xenopus laevis exposed to
aqueous leachates/percolates
Significant incr se in MN for leachate
of the PAH con minated soil
[88]
a Drosophila wing spot test [92,467].
d
M
r
n
ng
ha
tiv
M
s
ff
ff
e
m
li
ct
te
ea
ta
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 277
toxicity of organic fractions from a soil collected at a
highway junction that receives heavy vehicle traffic
[116]. The results revealed that fractions containing
polar aromatics such as nitroarenes induced significant
increases (�2-fold above control) in SCE frequency
both with and without S9.
8.2. In vivo eukaryote assays
Table 19 summarizes eight studies that employed in
vivo assays to assess the genotoxic activity of
contaminated soils. Three of these studies employed
the somatic mutation and recombination test
(SMART) in Drosophila melanogaster [68,203,321].
The assay, sometimes referred to as the wing spot test,
can detect both somatic mutations and recombination
in individuals that are trans-heterozygous for two wing
morphology mutations, multiple wing hairs (mwh) and
flare (flr) [92,322,323]. The study by Chroust et al.
exposed Drosophila eggs to DCM or water extracts of
soils collected in close proximity to a pharmaceutical
manufacturing plant and a coal tar conversion facility
[68]. Examination of adults raised from the exposed
eggs revealed significant increases in wing spot
frequency for DCM extracts of all four soils collected
near the coal-tar facility. Two of the water extracts
yielded weak positive responses. Additional chemical
analysis revealed that three of the four tested samples
had PAH levels above 700 ppb. Only two of the four
soils collected near the pharmaceutical manufacturing
facility yielded DCM extracts that elicited a positive
response in the wing spot assay. These results
corresponded to those obtained using the Arabidopsis
gene mutation assay and the SCE assay in Vicia faba.
DCM extracts of all four coal tar soils induced a
positive response in the Arabidopsis assay, and water
extracts induced significant increases in SCEs in Vicia
faba.
Two Russian studies used the SMART (wing spot)
to examine the genotoxicity of contaminated soils.
Gevirkian et al. examined the mutagenic activity of
organic extracts of soils collected from a swampy area
near a damaged waste pipe from a concrete production
facility in Khazakstan [321]. The results indicate no
significant increase in wing spot frequency over the
control. Pererva et al. employed the SMART to
examine the mutagenicity of extracts from 12
unidentified soil samples [203]. None of the samples
elicited a significant positive response. A third Russian
study employed the sex-linked recessive and dominant
lethal assays in Drosophila to examine the mutagenic
hazards of a radionuclide-contaminated soil [324].
The assays, first conceived in the late 1920s, can assess
the frequency of heritable (paternal) mutations, and
sequential mating can assess the susceptibility of
different germ cells stages (e.g., spermatagonia,
spermatocytes, spermatids) [325]. The results revealed
that a 14-h exposure of males to soils containing 7 Bq/
g significantly increased (�2-fold) the frequency of
sex-linked recessive lethals. Using a sequential
brooding pattern the dominant lethal assay revealed
that the maximum was associated with the pre-meiotic
spermatogonial stage.
Two studies employed an erythrocyte MN assay in
the clawed frog Xenopus laevis to assess the
mutagenic activity of contaminated industrial soils.
Chenon et al. exposed Xenopus larvae (i.e., tadpoles)
to aqueous extracts from soils amended with sewage
sludge [89]. Examination of blood after a 12-day
exposure failed to reveal a significant increase in MN
frequency. Bekaert et al. exposed Xenopus larvae to a
range of dilutions of aqueous extracts/leachates of two
soils, the first contaminated with solvents and metals,
the second contaminated with PAHs [88]. The results
revealed a significant increase in the frequency of
micronucleated erythrocytes (6–50%) in animals
treated with aqueous leachates of the PAH contami-
nated soil (�240 ppm). An aqueous percolate through
a column containing 30 kg of the metal contaminated
soil also elicited a significant increase in the frequency
of micronucleated blood cells (3%).
Two studies attempted to employ in vivo mamma-
lian assays to investigate the genotoxic hazards of a
contaminated soil. Wesp et al. employed the mouse
peripheral blood MN assay to assess the mutagenic
activity of various organic fractions from a soil
collected along a heavily trafficked motorway [116].
Gavage exposure of 7–12-week-old NMRI mice to
polar neutrals (e.g., trinitrotoluene), non-polar neu-
trals (PAHs), and polar aromatics (e.g., nitroarenes) at
2000 mg/kg followed failed to reveal any increase in
the frequency of micronucleated PCEs (polychromatic
erythrocytes) over the vehicle control. Bordelon et al.
fed Fisher 344 rats a diet that was supplemented with
coal-tar amended soil (0.35% by weight) for 17 days
and examined the frequency of bulky DNA adducts in
P.A
.W
hite,
L.D
.C
laxto
n/M
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8Table 20
Summary of soil mutagenicity results published only as conference abstracts
Site(s) examined Bioassay(s) employed Results obtained Reference
Soils from Tokyo, Bangkok,
Chaing Mai, and Manila
Salmonella mutagenicitya (TA100 and
TA98), benzene-EtOH (3:1) extract
Increased activity with S9. TA98 +S9:
Tokyo > Manila > Chaing Mai > Bangkok
[327]
Agricultural soils (Illinois, USA). Salmonella mutagenicity (preincubation
with TA98), water extract, combined MetOH–acetone
(1:1), MetOH–acetone–benzene (1:1:1) extract
Water extract: no activity. Organic extract
with S9: 3.5–5-fold increase over background
[468]
PAH contaminated soils (Czech Republic) Salmonella mutagenicity (TA100 and
TA98) and SOS Chromotest
With S9 activation: positive responses
on TA98, TA100 and SOS Chromotest
[328]
16 contaminated soils (PAHs, metals)
(Lumbardy region, Italy)
Salmonella mutagenicity, Tradescantia MNb
test, CAsc in Allium cepa root
Several positive responses. Details not provided [329]
Heavily contaminated (Superfund) soils (USA) Tradescantia MN test and CA test in Allium
cepa root tip cells on aqueous extracts
No response on Tradescantia MN test,
dose-related increases in CAs in Allium
[330]
Heavily contaminated (Superfund) soils (USA) CA test in Allium cepa root tip
cells on aqueous extracts
Dose-related increases in CAs in Allium [331]
Soil (ex situ) amended with
municipal wastewater sludge
Salmonella mutagenicity (TA98 with S9)
on aqueous leachate and hexane-acetone extract
Positive response on leachate and organic
extract of soil and sludge
[464]
Contaminated soils (not described) Tradescantia MN and SHM test, in situ incubations Not described [285]
Rubber factory mire (China) Tradescantia MN on mire extracts (solvent not specified) Significant increase in MN frequency above control [286]
Soils contaminated with heavy cycle oils Salmonella TA100 mutagenicity on
organic and aqueous extracts
Positive response on organic extract only [469]
Sewage sludge extracts applied
to native clay loam
Salmonella TA98 mutagenicity on DCM extracts Positive response without S9 [470]
Pesticide and wood preserving
waste contaminated soils
In situ Tradescantia MN and Zea
mays waxy locus assay
Significantly higher MN frequency than control.
Toxic effect inhibited maize growth
[332]
Street soils from metropolitan
Manila (Philippines)
rec� differential survival assay, MN assay
(organism not specified) on organic extracts
Significant mutagenicity and clastogenicity
with metabolic activation
[471]
Soils from rural, low density
traffic areas in Germany
Salmonella TA98 and TA100 mutagenicity
on hexane/acetone extracts
TA98 mutagenicity exceeded control for a variety
of samples. Lower TA100 mutagenicity. Response
not related to soil PAH concentration (0.03–2.6 ppm)
[472]
Urban soils (Japan) Salmonella mutagenicity (TA100, TA98,
TA98NR, TA98/1,8DNP6) on organic extracts
Highest activity in TA98NR and TA100 without S9 [473]
a Standard plate incorporation assay, unless otherwise noted.b Micronuclei.c Chromosomal aberrations.
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 279
lung and liver (dose �184 to 251 mg PAH/g rodent
chow) [326]. The results revealed a significant
increase in adduct frequency and a 3-fold greater
adduct frequency in lung versus liver. Moreover, no
declines in adduct frequency were seen in a group
exposed to coal tar amended soil that was aged for 9
months prior to dosing. Additional analyses showed
that several of the bulky adducts detected using 32P
post-labelling correspond to the metabolized products
of benzo[a]pyrene, benzo[b]fluoranthene, benzo[j]-
fluoranthene, and chrysene.
9. Miscellaneous soil genotoxicity results
The peer-reviewed literature also contains abstracts
of 20 studies that examined the genotoxic activity of
contaminated soils and soil extracts. Table 20 contains
a summary of these studies. Of the 20 studies listed,
the majority (9) employed the Salmonella mutageni-
city test, 5 used the Tradescantia MN test, 3 assessed
anaphase aberrations in Allium root tip cells, one
examined waxy locus mutations in Zea mays, one
examined differential survival in rec� B. subtilus, and
one employed the SOS Chromotest. Despite a lack of
detailed information about these studies and the
results obtained, the table does contain some inter-
esting and useful information. For example, the
Salmonella mutagenicity results obtained for extracts
of urban soils [327], and soils known to be
contaminated with PAHs [328,329] are consistent
with the results already discussed (i.e., S9-activated
positive responses in TA100 and TA98 accompanies
PAH contamination).
Two studies that employed the Tradescantia MN
assay and the Allium anaphase aberration assay to
examine aqueous extracts of heavily contaminated
Superfund soils revealed a lack of response in the
Tradescantia assay and a significant positive on the
Allium assay [330,331]. This result is consistent with
the Tradescantia results already discussed in Section
8.5 (i.e., Tradescantia MN assay is less responsive to
aqueous extracts). In addition, two of the listed studies
did not detect significant induction of MN in
Tradescantia tetrads following direct contact expo-
sures [285,332], or exposures to soil organic extracts
[286]. This is also consistent with the results presented
in Section 8.5.
10. Remediation of genotoxic soils
Contaminated soils can be remediated using a
variety of techniques that fall into four basic
categories: physical, chemical, thermal, and biological
[333]. Physical techniques include a variety of
methods for excavation, entombment, or covering of
contaminated areas [333,334]. Excavation involves
removal for on- or off-site destruction, disposal or
storage. Entombment involves construction of a
properly sealed landfill used to retain highly con-
taminated soils and prevent movement of toxic
contaminants. Covering, a method that is not
acceptable for highly contaminated areas, simply
involves covering the contaminated material with
fresh soil, concrete, or asphalt.
Chemical methods include a wide range of techni-
ques to neutralize or remove toxic substances. Common
strategies include soil washing, vacuum extraction,
electro-reclamation for metal removal, contaminant
oxidation, precipitation, or reduction, and contaminant
adsorption to activated carbon [335–338]. Thermal
techniques essentially involve detoxification via com-
bustion. Commonly used techniques include fluidized
bed or rotary kiln combustion [333,337].
Biological remediation techniques involve the use
of organisms (e.g., bacteria, fungi, plants) to detoxify
and/or destroy toxic contaminants. The strategy is
mainly applied to soils contaminated with organic
materials and a wide range of techniques are currently
available [333,337–344]. The popular bioremediation
techniques can essentially be sub-divided into four ex
situ categories: biopile, bioslurry, composting, and land
farming; and three in situ categories: attenuated
bioremediation, passive bioremediation, and phytor-
emediation [333, 337,338,340–343,345–347]. Biopile
treatment involves excavation, piling the contaminated
material in mounds, and incubating the material under
natural conditions for extended periods of time [348–
350]. The method usually involves supplementation
with air, and sometimes nutrients, minerals, and water
are added to speed degradation [349,350]. In addition,
organisms selected for their ability to degrade particular
contaminants (e.g., petroleum hydrocarbons) are some-
times added [348]. Composting, a method that is similar
to biopiling, involves supplementation of the soil with
manure (e.g., bovine, equine) and/or dry plant material
(e.g., agricultural waste, sawdust) to provide a source of
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345280
organic carbon and enhance the concentration of
nutrients such as phosphorus and nitrogen
[66,188,350–352]. The strategy is designed to provide
temperature, moisture content, aeration, and nutrient
concentrations that promote degradation. The bioslurry
technique is a controlled, sophisticated variation on the
biopile technique. Treatment by bioslurry involves
mixing the soil with water, nutrients, and air and
incubating the resulting slurry in a bioreactor under
controlled conditions [44]. The incubation temperature
is controlled, and the mixture is often continually
supplemented with nutrients and air. The technique also
frequently involves the addition of microbes selected
for their ability to degrade selected contaminants
[353,354]. Land treatment or land farming, a technique
that is fairly similar to biopiling, involves tilling
of the contaminated soil with nearby uncontaminated
soils, followed by incubation and monitoring for
an extended period of time. The material is usually
tilled and irrigated at some predetermined frequency
to enhance degradation by indigenous organisms
[44,115,355,356]. In areas with marginal or acidic
soils the tilled mixture is often amended with buffering
agents and/or nutrients.
In situ bioremediation techniques involve degrada-
tion of soil and/or groundwater contaminants by the
native fauna, or augmented native fauna, without site
excavation. A popular and effective technique, known
as accelerated natural attenuation (ANA), involves
injection of compounds that accelerate the degrada-
tion of toxic contaminants by natural microbial flora.
For example, researchers have developed, and are now
successfully marketing, compounds that release
oxygen for accelerated in situ aerobic degradation,
or hydrogen for accelerated in situ anaerobic
degradation [357–359]. Two products developed to
enhance in situ aerobic or anaerobic remediation,
ORC1 (oxygen releasing compound) and HRC1
(hydrogen releasing compound), can be pressure
injected into the contaminated area for ANA (see
www.regenesis.com/products). This strategy is parti-
cularly effective for stimulation of reductive dechlor-
ination and remediation of sites contaminated with
chlorinated solvents such as tetrachloroethene (also
known as tetrachloroethylene or perchloroethylene)
[357]. In addition to supplementation with oxygen-
and hydrogen-releasing compounds, ANA can also
involve introduction of organisms selected for their
ability to degrade particular substances. Recently
isolated bacteria belonging to the genus Dehaloco-
coides effectively accelerate the degradation of
trichlro- and tetrachloroethene, generating only
ethene, biomass, and inorganic chloride [360,361].
The term phytoremediation encompasses a wide
range of techniques that employ plants to extract,
degrade, transform, or sequester toxic contaminants in
soils. Various terms have been employed to describe
specific applications and strategies used in phytor-
emediation. These include phytoextraction, phytode-
gradation, phytotransformation, phytovolatilization,
and rhizodegradation [340]. Phytoextraction involves
the use of carefully selected plants to accumulate toxic
contaminants (e.g., metals) in above ground biomass.
The plants are then harvested, incinerated, and the
ashes properly disposed [362,363]. Phytodegradation
and phytotransformation involve uptake of soil
pollutants and metabolic transformation into less
toxic or non-toxic compounds [364]. A specific form
of phytotransformation known as phytovolatilization,
involves atmospheric release of toxic contaminants or
their metabolites via plant transpiration [340,365].
Rhizodegradation, a variation of the same theme,
involves degradation in the microbe-rich zone around
the root system of a vascular plant (i.e., the rhizo-
sphere). This technique has been successfully applied
to soils contaminated with PAHs and petroleum
hydrocarbons [366].
A detailed overview of the remediation technologies
used at sites contaminated with mutagenic substances is
clearly beyond the scope of this work. For detailed
information on the various soil remediation technolo-
gies readers can consult Terry and Banuelos [367],
Suthersan [368], Sara [369], Singh and Jain [340], Sims
[337] or Ritter and Scarborough [339].
Table 21 summarizes the results of published
studies that investigated the ability to remediate soils
contaminated with genotoxic substances. The table
summarizes the results of 30 remediation assessments
from 26 studies. Six assessments examined soils
contaminated with petrochemical wastes (e.g., slop-
oil emulsion, oil–water separator sludge) or petroleum
distillates (e.g., diesel oil, used motor oil) [59,64,121,
154,157,166]. Nine assessments examined soils
contaminated with wood preservation wastes or
creosote [44,56,64,115,120,121,177,370]. Five stu-
dies examined soils contaminated with PAH-rich
P.A
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tion
Resea
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1
Table 21
Summary of studies investigating the (bio)remediation of mutagen-contaminated soils
Sample/site Contamination Assay(s) employed Remediation process Treatment
time (days)
Results obtained Reference
Soil (sandy loam) amended with
petrochemical wastes (slop
oil emulsion)
PAHsa (�6000 ppm), oil and
grease, heavy hydrocarbons
Salmonella TA98 mutagenicity on
DCMb extracts
Batch reactor degradation.
Water content adjusted.
Temperature = 20 8C
354 Mutagenicity (+S9 and �S9) reduced
to below detection. 43% reduction in
PAH concentration
[121]
Soil (sandy clay loam) amended
with API oil–water separator sludge
Heavy hydrocarbons, PAHs Salmonella TA98 mutagenicity on
DCM and MetOH extracts
Incubation in barrel lysimetersc
exposed to normal rainfall
350 Slight increase in mutagenicity (+S9)
at 180 days. Decrease to background
by 350 days
[64]
Soils (sandy clay and clay) amended
with petrochemical wastes (SWRI
wasted and slop-oil emulsion solids)
Petroleum hydrocarbons, PAHs Salmonella TA98 mutagenicity and
forward mutation induction in
Aspergillus nidulans (DCM extracts)
Incubation in wooden boxes.
Periodic moisture adjustment
360–1000 Steady decline in mutagenicity (+S9)
over time
[166]
Soil (clay loam) amended with
petrochemical sludge
PAHs (473–3782 ppm) Salmonella TA98 on ACN extracts Laboratory bioremediation.
Adjusted moisture and temperature
373 Marked decline in �S9 response. Initial
increase in +S9 response followed by
slower decline
[59]
Soil contaminated with
used motor oil
Petroleum hydrocarbons, PAHs Salmonella TA1535 and TA1537
mutagenicity on aqueous extracts
Inoculation with bacteria and
incubation in plastic containers
at 25 8C. Moisture adjusted and
nutrients added
84 Steady decline in mutagenicity to
below detection (�S9)
[154]
Soil (sandy loam) amended
with diesel oil
Diesel oil (PAHs) Salmonella TA98 mutagenicity
on DCM extracts
Bioremediation (liming, fertilization,
and tilling) in outdoor lysimeters
�140 Bioremediation treatment eliminated
mutagenicity (�S9) and PAHs
in 12 weeks
[157]
Soils collected near a coke
production facility
PAHs (600–800 ppm). SOS Chromotest, Salmonella TA98
mutagenicity on organic extracts
Unspecified biodegradation process 210 Large reductions in mutagenicity
(�S9 and +S9) and PAH contamination
[153]
Composted soil from a former
gas-works site (Czech Republic)
PAHs (610 ppm in unaltered
soil), metals
SOS Chromotest on DCM extract 4 to 1 (wet weight) mixture of
compost (wheat straw, chicken
manure, gypsum) and soil
54 Slight reduction in genotoxicity
(+S9) in upper portions of composted
pile. 20–60% reduction in PAHs
[188]
Soil (Lima loam) amended with
coal tar extract
PAHs (400–1400 ppm). Rfpe in Pseudomonas putida (in situ) Biodegradation in situ
(land treatment) and ex situ
(laboratory)
59–180 Marked decline in PAH concentration
and mutagenicity by 147 days. PAH
reduction not related to genotoxicity
changes
[90]
Soil near coke oven work (Portugal) PAHs (1141 ppm) Mutatox1 assayf on aqueous eluates Environmentally controlled
landfarming greenhouse
150 60% reduction in total PAHs. 80%
reduction in 2-, 3-, and 4-ring PAHs.
Weak mutagenicity in eluate from
untreated sample only
[201]
Coal tar contaminated soil Variety of 3-, 4-, 5-, and
6-ring PAHs
SOS Chromotest on aqueous leachate Incubation at 30 8C in 1 L
stirred (glass) reactors
50–90 No genotoxicity detected (�S9 and
+S9). 20–70% reduction in
concentration of mutagenic PAHs
[189]
Soil (sandy loam) amended with
wood preserving wastes
(creosote sludge)
PCPg (�2500 ppm), creosoteh,
PAHs, dioxins
Salmonella TA98 mutagenicity
on DCM extracts
Batch reactor degradation.
Water content adjusted.
Temperature = 208C
354 Mutagenicity (+S9) reduced to
below detection. 71% reduction
in PAH concentration
[121]
Contaminated soil (loam or sandy
loam) from a wood-preserving
facility
PCP (31–176 ppm), carcinogenic
PAHs (126–279 ppm), Cu, Cr,
and As
Salmonella TA98 mutagenicity
on DCM and MetOH extracts
Bioremediation (tilling,
inoculation, and nutrient
addition) in an LTUi
�90 Mutagenicity (+S9) of most samples
reduced to acceptable levels
(<150 rev/g) within 3 months
[115]
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
28
2Table 21 (Continued )
Sample/site Contamination Assay(s) employed Remediation process Treatment
time (days)
Results obtained Reference
Contaminated soil from a
wood-preserving facility
Creosote, Zn, PAHs, dioxins Salmonella TA98 mutagenicity
on DCM extracts
Bioremediation in an LTU
(12 separate cells)
�240 Mutagenicity (+S9) increased in
nine of 12 LTU cells. Six
increases >2-fold
[56]
Soil (clay loam) amended
with wood-preserving waste sludge
PCP, creosote, PAHs, dioxins Salmonella TA98 mutagenicity
on DCM/MetOH extracts
Incubation in barrel lysimeters 350 Increase in mutagenicity to 180 days,
following by a decline below initial
for +S9 only
[370]
Soil (sandy clay loam) amended with
sludge from a wood-preserving
plant impoundment
Creosote, PCP, PAHs, dioxins Salmonella TA98 mutagenicity
on DCM and MetOH extracts
Incubation in barrel lysimeters
exposed to normal rainfall
350 Increase in mutagenicity at 180 days
(�S9 and +S9). Some decline
by 350 days
[64]
Creosote contaminated soil PCP, PAHs, creosote, dioxins Salmonella TA98, TA100, TA97,
TA102, TA104, YG1042 and
YG1042 mutagenicity on DCM extracts
Biopilej, bioslurryk, compostl,
and land treatmentm140, 41, 84,
and 175
Large increase (�S9 and +S9) in
bioslurry sample. Moderate increase
(�S9 and +S9) in biopile sample
[44]
Creosote contaminated soil PCP, PAHs (>5000 ppm),
dioxins
Tradescantia MN test on aqueous extracts Fungal inoculation and
simulated land treatment
56 Remediation resulted in reductions
in soil PAH content and soil
clastogenic activity
[120]
Wood preserving bottom
sediment (EPA K001)
PCP, creosote Salmonella TA98 mutagenicity on
DCM/MetOH extracts
Potassium polyethylene glycol
treatment (KPEG) at 38, 48, 80
and 120 8C
<1 Slight increase in mutagenicity
(+S9) followed by rapid decline
within 30 min
[177]
Wood preserving bottom
sediment (EPA K001)
PCP, creosote E. coli prophage induction assay on
DCM/MetOH extracts
Potassium polyethylene glycol
treatment (KPEG) at 38, 48, 80
and 120 8C
<1 High toxicity, difficult to interpret
and determined effects of remediation
[177]
Soil (fine loam) amended with
municipal wastewater sludge
(activated)
Unknown Salmonella TA98 and TA100
mutagenicity on ethylene
dichloride extracts
Eight week incubation with
periodic additions of water
56 Increase in mutagenicity (�S9)
for the first 7 days, decline to
background after 21–42 days
[372]
Soil (sandy clay and sandy loam)
amended with sewage sludge
Unknown Salmonella TA98 mutagenicity on
DCM and/or MetOH extracts
Incubation in undisturbed
barrel lysimeters
717 Some increase in mutagenicity
(+S9) during the first �40 days.
Decrease (�S9 and +S9) to
background by 700 days
[179]
Soils (sandy clay and sandy loam)
amended with sewage sludge.
Unknown Salmonella TA98 mutagenicity on
DCM and MetOH extracts
Incubation in undisturbed
barrel lysimeters
510 Increase in mutagenicity (+S9 only)
to 154 days, followed by slow
decline to background by 510 days
[156]
Soils amended with municipal
wastewater
Unknown Salmonella TA98 mutagenicity on
DCM extracts
Soils amended with wastewater
extracts incubated at room
temperature
2 Some decline in mutagenicity
(�S9) within two days
[175]
PCBn contaminated soil Aroclor 1260 (�2200 ppm) Salmonella TA98 and TA100
mutagenicity on DCM extracts
Base-catalyzed dechlorination
at 250–350 8C<1 50% reduction in TA98
mutagenicity (�S9 and +S9)
[41]
PCB contaminated soil Aroclor 1260 (�2200 ppm) E. coli prophage induction assay
on DCM extracts
Base-catalyzed dechlorination
at 250–350 8C<1 No appreciable reduction in
genotoxic activity (�S9 and +S9)
[41]
PCB contaminated soil Aroclor 1260 (�150 ppm),
PAHs, dioxins
CAs in Allium cepa Pilot-scale solvent extraction <1 99% reduction in PCBs. Increase
in clastogenicity and phytotoxicity
[254]
Soils contaminated with
solvent-recovery wastes and
paint sludge
Xylene, toluene, heavy metals Salmonella TA98 mutagenicity on
DCM and MetOH extracts
Physical removal of visible
contamination
Unknown Dramatic reductions in soil
mutagenicity (�S9 and +S9),
but levels remained �30-fold
background
[114]
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 283E
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wastes such as coal tar or coke oven emissions
[153,188,189,201,371]. Four studies examined soils
amended with sewage sludge or municipal waste-
waters [156,175,179,372]. Three assessments in two
studies examined PCB-contaminated soils, two stu-
dies examined soils contaminated with munitions
waste or explosives [158,200], and the final study
examined a soil contaminated with solvent recovery
wastes and paint sludge [114].
The first six entries in Table 21 suggest that the
mutagenic hazards associated with soils containing
petrochemical wastes or petroleum distillates such as
diesel oil and motor oil can be effectively remediated.
The two studies that employed bioremediation with
nutrient supplementation for soil contaminated with
motor oil or diesel oil showed relatively rapid declines
in mutagenicity to below detection within 7–12 weeks
[154,157]. The studies that examined soils contami-
nated with more complex petrochemical wastes
containing an assortment of heavy hydrocarbons
and aromatic substances showed effective remediation
within one year [59,64,121,166]. It is interesting to
note that one study that followed petroleum waste
amended soils incubated in barrel lysimeters (i.e., soil
monoliths), noted a moderate increase (i.e., 1.8-fold)
in S9-activated mutagenicity during the first 180 days
that may be due to the liberation or formation of
mutagenic compounds during the remediation process
[64]. Although this study did not investigate the
accumulation of any mutagenic degradation by-
products or the persistence of any contaminants, a
recent bioslurry study of a PAH-contaminated soil
from a former gasworks site noted the persistence of
alky-PAHs and N-heterocyclics, as well as an increase
in the concentration of oxygen-containing PAHs over
the course of the remediation [353].
Five of the listed studies investigated the ability to
remediate the genotoxic hazard associated with PAH-
containing wastes such as coal tar or coke oven
emissions [90,153,188,189,201]. The results obtained
in these studies indicate that large reductions in the
concentrations of 3- and 4-ring PAHs can be achieved
within 200 days. Two studies observed 20–70%
reductions in PAH concentrations within 50–100 days.
However, a reduction in genotoxic activity did not
always correspond to the noted reductions in PAH
concentration. For example, compost remediation of
PAH-contaminated soils (>600 ppm) from a former
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345284
gas-works site resulted in a 20–60% reduction in PAHs
in 54 days, but very little reduction in the SOS
genotoxicity of a DCM soil extract [188]. However, two
similar studies that examined a soil collected near coke
production facility (PAHs �600–800 ppm), and a soil
amended with coal tar extract (PAHs�400–1400 ppm),
both noted large reductions in both mutagenicity and
PAH concentration within 147 and 210 days, respec-
tively [90,153]. In addition, the latter study observed a
lack of relationship between PAH concentration decline
and the decline in mutagenicity (i.e., induction of
rifampicin resistance in Pseudomonas putida) [90]. The
seeming incongruence of these results is likely due to
wide variations in the efficacy of bioremediation for the
removal of PAHs and mutagenic activity from soils
contaminated with coal tar and coke oven emissions.
Several studies have demonstrated that the efficacy of
PAH removal via bioremediation is heavily influenced
by PAH composition, the selected bioremediation
process, the physical, chemical, and biological proper-
ties of the soil, and the environmental conditions during
remediation [341,373,374]. Moreover, cross-study
comparisons of mutagenicity values recorded before,
during and after the aforementioned remediations is
complicated by variations in extraction solvent and
endpoint selection.
Eight studies investigated the ability of bioremedia-
tion to remove the genotoxic hazards associated with
soils contaminated by wood preserving wastes. It is
interesting to note that four of these studies recorded
substantial increases in mutagenic activity during the
remediation [44,56,64,370]. For example, the study by
Hughes et al. of a creosote contaminated soil from the
Reilly Tar Site noted that DCM extracts failed to elicit
significant positive responses on Salmonella strains
TA98, YG1041 and YG1042, and elicited only a weak
response on TA100 (without S9) [44]. However, after
bioslurry remediation, DCM extracts elicited signifi-
cant positive responses on YG1041 and YG1042, as
well as TA98 with S9. Additional positive responses
were observed following biopile remediation (YG1041
with and without S9, YG1041 with S9, TA100 with and
without S9) and composting (YG1041, YG1042, TA98
and TA100 without S9). Additional analyses of the
bioslurry sample indicated that the putative mutagens
included nitroarenes. Studies by Barbee et al. showed a
moderate increase in S9-activated TA98 mutagenicity
and a substantial increase in direct-acting TA98
mutagenicity after 180 days in a barrel lysimeter
exposed to normal rainfall [64,370]. Although this was
followed by a decline in mutagenic activity, the direct-
acting activity after 350 days was still substantially
above that recorded before remediation. Neither the
Hughes et al. nor the Barbee et al. studies determined
the source(s) of the increased mutagenic activity.
However, Hughes et al. suggested that materials (e.g.,
nutrients, manure, plant biomass) added to the
bioslurry, biopile, and compost mixtures may have
contributed mutagenic compounds. This assertion is not
supported by other studies, such as the works of
Kazunga et al. and Lundstedt et al., which noted that
bioremediation of PAH-contaminated soils can result in
the formation of oxygen containing PAHs and these
compounds can accumulate over the course of the
remediation [353,375]. Some of these compounds, such
as a variety ortho-quinones and ketones, are known to
be potent direct-acting Salmonella mutagens (TA98)
[376–378], as well as mammalian cell mutagens (tk
locus in human lymphoblastoid cells expressing several
P450 enzymes) [379].
It is interesting to note that two additional studies on
soils contaminated with wood-preserving wastes
observed substantial declines in mutagenic activity to
levels deemed to be acceptable (e.g., <0.15 reverants/
mg) within as little as 90 days [115,121]. For example,
Donnelly et al. noted that bioremediation in a land
treatment unit supplemented with nutrient and lyophi-
lized bacteria resulted in substantial mutagenicity
reduction within 3 months [115]. April et al. noted
that batch reactor incubation of soils amended with
wood-preserving wastes for 354 days resulted in a
decline in Salmonella mutagenicity to below detection
[121]. An aggressive chemical remediation method
employing potassium polyethylene glycol (KPEG) at
temperatures up to 120 8C was employed by Hong et al.
to remediate a wood preservation site impoundment
sediment [177]. The results revealed substantial
reductions in PCP concentration (>80%) within
30 min, and corresponding reductions in S9-activated
TA98 mutagenic activity.
One study by Donnelly et al. investigated the
mutagenic potential of soils collected before and after
remediation of an abandoned solvent recovery site
[114]. Visual inspection of the remediated site indi-
cated that all visible signs of contamination had
been removed; moreover, the TA98 mutagenic activity
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 285
of DCM/MetOH extracts showed a marked reduction.
However, extracts of soils collected from a runoff ditch
still exhibited mutagenic activity up to 30-fold above
background.
Four studies investigated the ability to remediate
mutagenic soils that have been amended with
municipal waste water or sewage sludge [156,175,
179,372]. Three of these studies noted an initial, but
short-lived (i.e., 7–154 days) increase in mutagenic
activity, followed by a marked decline to background.
The fourth study only followed the remediation for
two days, but did note some decline in mutagenicity
[175,372]. The time required to reach background
mutagenic activity varied between the three afore-
mentioned studies. Angle and Baudler noted an
increase in the direct-acting TA98 and TA100
mutagenic activity of extracts from soil amended
with sewage sludge, followed by decline to back-
ground levels within 21–42 days [372]. In two similar
studies, Donnelly et al. noted initial increases in the
S9-activated TA98 mutagenicity of sludge-amended
soils during the first 50–150 days in an undisturbed
lysimeter [156,179]. This was followed by a decline to
background (TA98 with and without S9) within 510 to
770 days. Thus, the time required for complete
removal of mutagenicity could be as long as two years.
Two studies employed aggressive extraction or
thermal degradation strategies to remediate the
mutagenic activity and PCB content of contaminated
soils [41,254]. DeMarini et al. demonstrated that base-
catalyzed dechlorination of PCB-contaminated soils at
250–350 8C (�8 h) removed 99% of the PCB
contamination and reduced the TA98 mutagenic
activity (with and without S9) by approximately 50%
[41]. However, the same study noted that the base-
catalyzed dechlorination process did not reduce the
genotoxic activity measured using the prophage
induction assay. Meier et al. demonstrated that a
pilot-scale soil washing procedure effectively removed
PCB contamination from soils collected from a former
transformer storage site [254]. However, remediation
resulted in an increase in the clastogenic activity as
measured using the Allium anaphase aberration assay.
This increase was removed following a water rinse step,
and the authors suggest that residual wash solvent
caused the initial increase in clastogenic activity.
Although explosives and munitions wastes (e.g.,
TNT, RDX, HMX, hexogen) are noted for their
mutagenic activity, toxicity and environmental per-
sistence, few studies have attempted to investigate the
ability to remediate the mutagenic hazards of
munitions contaminated soils. Two of the studies
listed in Table 21 used composting to remediate the
mutagenic hazards of soils contaminated with a
variety of munitions and explosive residues. However,
the two studies reached opposite conclusions. The
study by Jarvis et al. observed marked increases in
direct-acting mutagenicity (Mutatox1) following 30-
day composting (20% soil by weight) in an adiabatic
reactor [200]. Simultaneous chemical analysis noted
marked reductions in TNT, RDX, 2-amino-DNT and
4-amino-DNT. The authors suggest that degradation
of the explosives may have resulted in the formation of
mutagenic contaminants that were not detected using
routine chemical analyses. The study by Robidoux et
al. (see Table 11) also had difficulty reconciling the
results of chemical analysis and the direct-acting
genotoxic activity (SOS Chromotest) of elutriates
from munitions contaminated soils [193]. In contrast,
the study by Griest et al. noted substantial decreases in
the TA98 and TA100 direct-acting mutagenicity in
ACN extracts of soils remediated by static or
mechanically stirred composting for 44 or 90 days,
respectively (10% soil by weight) [158]. Simultaneous
chemical analysis showed that the TA98 and TA100
direct-acting mutagenic activity at the start of the
experiment (i.e., compost day 0) was consistent with
the measured concentration of TNT, and moreover,
that the decline in mutagenicity was accompanied by a
corresponding decline in TNT concentration. The
seemingly incompatible outcomes of these two studies
may be related to oxygen concentration, the soil
concentration in the compost, and the composition of
the microbial community in the compost pile. Several
researchers have previously noted large variations in
the ability of microbial assemblages to degrade
explosives such as TNT, RDX and HMX, as well as
increased mineralization rates in the absence of
oxygen [346,354].
11. Summary and discussion
A wide range of human activities (e.g., industrial,
municipal, rural, urban) has contributed to the
contamination of land. Industrial practices including
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345286
improper use and storage of hazardous materials,
wastewater discharge, waste disposal, waste storage,
and accidental spills or leaks have resulted in serious
contamination problems in industrialized nations. Use
of industrial and municipal wastewaters for irrigation
and heavy use of herbicides, fungicides and insecticides
have contributed to contamination in rural, agricultural
regions. In addition, combinations of activities,
including fossil fuel combustion and surface deposition
of combustion by-products, solid waste disposal, and
waste incineration, have contributed to the contamina-
tion of soils in urban areas. The fate of these substances
depends on complex dynamic processes that are
controlled by environmental conditions (e.g., climate),
soil physical properties (e.g., texture, water content,
cation exchange capacity, organic matter content), soil
biological properties (e.g., microbial community
structure), and the physical-chemical properties of
the contaminants (e.g., Kow, vapour pressure, specia-
tion, charge, stability). Persistent, insoluble organic
compounds are virtually immobile in soil and will
remain in place for an extended period of time (i.e.,
years to decades) [5,7]. Although the mobility of these
compounds will be partly determined by water move-
ment, soil porosity, and soil organic matter content,
organic compounds with log Kow values greater than
4.0 (e.g., PCBs, PCP, BaP) are generally considered to
be immobile [5]. Substances with log Kow values less
than 2.5 (e.g., tetrachloroethane, benzene) are often
highly mobile and can move to the water-saturated zone
with relative ease. Stable organics with high vapour
pressures and low water solubility (i.e., high Henry’s
law constant Kh) will tend to volatilize, move through
soil pores, and exit the soil as a vapour [5,7]. The
mobility of toxic metals (e.g., Pb, Cd, Cr, Zn, Sb, Ni,
Hg) is a complex function of soil redox (Eh), soil
texture and cation exchange capacity (e.g., clay
content), soil pH, and metal speciation. As a general
rule cations, including many divalent metals that are
highly toxic, have low mobility in silt and clay soils,
and moderate mobility in sandy soils [5,7].
The risk of adverse health effects attributable to
contaminants in soils will be determined by environ-
mental fate, exposure, and toxicity. Substances that are
rapidly mineralized will present little opportunity for
exposure and adverse effect. The route and magnitude
of exposure to toxic substances in soil depends on the
properties of the substance, as well as the life history
and physiology of the organism under consideration.
Exposures to volatile substances generally occur via
inhalation and/or dermal absorption [380–384], and
exposures to mobile organics and metals commonly
occurs through the ingestion of contaminated ground-
water [380,385–387]. Exposures to persistent immo-
bile contaminants that are strongly adhered to soil
particles, including many mutagenic substances (e.g.,
PAHs, creosote, petrochemical waste), can occur via
inhalation, non-dietary ingestion of soil particles
[388–392], or ingestion of contaminated biota [393].
Current risk assessment guidelines employ standard
residential, non-dietary soil ingestion values of
100 mg/day for adults and 200 mg/day for children
[391,394]. For commercial/institutional or outdoor
industrial exposures of adults, 50 mg/day and 480 mg/
day are used, respectively [394]. Although direct
exposure via the consumption of contaminated soil
organisms seems unlikely, wildlife and humans can be
exposed to soil contaminants via the consumption of
terrestrial biota which accumulate persistent soil
contaminants (e.g., arctic caribou) [393,395,396].
Moreover, humans can be exposed via consumption
of produce grown in contaminated soils. Several
studies examining animals (e.g., pigs, sheep, quail) fed
grain grown on land amended with sewage sludge
showed elevated organ (e.g., kidney, liver) levels of Cd
and Ni [397–399]. In addition, Telford et al. showed
that sheep fed sugar beets grown on sludge amended
soils excreted mutagenic metabolites via the bile and
urine [400], and Shane et al. showed that the tissues of
lettuce and endives grown in flyash-amended soils
contained S9-activated TA98 mutagens [401].
Volatile contaminants in soil can constitute an
extreme hazard if environmental conditions permit
sufficient exposure via air or water. The most familiar
scenario whereby a volatile soil constituent can
enhance the risk of mutagenic and carcinogenic
effects is radon gas exposure. Radioactive isotopes of
radon, a by-product of the 238U decay series, can seep
into buildings and dramatically enhance the risk of
mutagenic effects and lung cancer [402,403]. Beyond
radon, there are very few well-documented, non-
occupational accounts of exposure and adverse effects
attributable to non-radioactive, volatile pollutants in
soils. Although several studies have documented
adverse health effects in individuals inhabiting regions
in close proximity to hazardous waste dump sites who
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 287
have complained of household odours and ill health
(e.g., respiratory distress, eye and skin irritations,
gastrointestinal disorders, cancer, birth anomalies,
neurological effects), connections, either empirical or
otherwise, to the presumed source of contamination
are tenuous [384,404–407].
Mobile metals and organic contaminants in soils
can readily contaminate groundwaters, and there are
well documented accounts of serious adverse health
effects (e.g., cancer, neurological defects, skin
disorders, reproductive problems) in communities
exposed to groundwaters that have become contami-
nated via nearby industrial activities and associated
soil contamination problems [80,384,385,387,405,
408,409]. Exposures generally occur via ingestion
of contaminated groundwater; however, dermal
absorption and inhalation during bathing and shower-
ing have recently been noted as potential routes of
exposure [384]. Nevertheless, although the connec-
tions between exposures to mobile soil contaminants
(i.e., exposure via groundwater) and adverse health
effects are more convincing, empirical connections are
often weak and mechanistic connections are lacking
[410–413]. One noteworthy exception is the well-
documented link between a variety of serious adverse
health effects, including cancer, neurological dis-
orders and skin lesions, and exposure to arsenic in
contaminated groundwater [414].
Many known mutagens are expected to be strongly
adsorbed to soil particulate matter. Thus, adverse
health effects attributable to these substances would
require exposures to the actual soil particles via
ingestion or inhalation. Although the aforementioned
risk assessment guidelines do acknowledge non-
dietary ingestion as a well-documented route of soil
exposure in commercial, residential, and industrial
settings [394,415], it is difficult to determine the
likelihood of human exposure to soil particles from a
contaminated area. Consequently, it is difficult to
accurately calculate the mutagenic risks associated
with contaminated soils, or habitation in close
proximity to a contaminated site. Nevertheless, three
studies by Najem et al. investigating the suggested
excess cancer mortality in the state of New Jersey
revealed that for several cancers, including breast and
gastrointestinal, there is a statistically significant
positive correlation between cancer mortality and the
distribution of hazardous chemical waste disposal
sites (HCWDS) [416–418]. A similar study by
Budnick et al. concluded that although bladder cancer
mortality rates in white males from Clinton County,
Pennsylvania, a county contaminated with mutagenic
carcinogens from a Superfund site, are elevated, there
is no consistent pattern of environmental exposure and
‘‘causal inferences cannot be made’’ [407]. In
addition, a study by Vine et al. noted that although
40–59-year-old residents living within one mile of a
pesticide dump site in North Carolina had increased
levels of serum DDE, residential location was ‘‘not
consistently associated’’ with the frequency of MN in
peripheral blood lymphocytes [419].
Despite this difficulty in quantifying the risk of
adverse health effects attributable to soil contamina-
tion, a great deal of effort has been expended to assess
site contamination, estimate risk, and remediate
contaminated sites. Indeed, this work is a review of
1312 assessments of soil genotoxic hazard. Although
some of the reviewed studies were indeed motivated
by well-documented risks associated with mobile
contaminants, most studies appear to have been
motivated solely by presumed risk and the associated
liability. The US National Oil and Hazardous
Substances Pollution Contingency Plan, the regulation
that implements CERCLA and establishes the
approach for determining potential health risks and
appropriate remedial action, calculates risks asso-
ciated with exposures of on-site workers in direct
contact with contaminated soils, current (nearby)
residents exposed to the site via contaminated air (i.e.,
volatiles and soil particles) and water, future residents
in direct contact with the site, and even future residents
exposed via consumption of contaminated fish from
on-site water bodies [420]. The comprehensive nature
of the risk assessments required under CERCLA and,
the liability for injury and loss of natural resources set
forth in section 9607(a) of Title 42 US Code Chapter
103, i.e., that the ‘‘owner and operator of the vessel or
facility causing the release of a toxic substances is
liable’’, rationalizes a cautious approach to site
assessment and site management [421]. It is this
cautious approach, as well as the desire to recover
contaminated land that may have substantial value,
that has encouraged and supported assessments of soil
genotoxicity.
An evaluation of the stated objectives in the 118
publications examined for this review reveals that
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345288
most of the studies were indeed motivated by a
requirement to assess the genotoxic hazards of current
contaminated sites, determine the mobility of the
genotoxic contamination, and evaluate the efficacy of
current remediation efforts. For example, of the 19
works published by KC Donnelly and colleagues at
Texas A&M University, 17 of the papers described site
assessment, investigations of genotoxicity mobility,
investigations of remediation efficacy, and evaluations
of current waste treatment practices [40,42,47–49,54,
56,64,114,115,117,118,
146,156,165,166,179,317,370] as the study objective.
The remaining two papers constituted a comparative
evaluation of short-term microbial assays for the
examination of soil genotoxicity [146,317]. In
general, over 45% of the publications examined for
this review appeared to be primarily motivated by a
requirement to assess potential genotoxic hazard. With
several studies also investigating the leachability of
genotoxic material [48,61,174,283]. This is quite
distinct from the situation encountered in a recent
review of the sediment mutagenicity literature. The
review by Chen and White [422] noted that most
studies assessing the mutagenic activity of contami-
nated aquatic sediments are motivated by an
abnormally high frequency of idiopathic lesions,
including hepatocellular carcinoma, in indigenous
fishes. Thus, the documented effect had already been
observed, and the motivating factor behind sediment
examinations was disease etiology.
The data collected for this review clearly indicates
that soils at most contaminated industrial sites, as well
as urban sites, are genotoxic and an enhanced risk of
mutagenic effect would likely accompany exposure to
these soils. Of course, the magnitude of the risk will
depend on the potency of the contaminated soil, as
well as the magnitude and route of exposure. The
collected Salmonella data presented in Fig. 3 and
Table 8 indicate that the mutagenic hazard (i.e., TA98
with and without S9, TA100 with S9) associated with
soils from contaminated industrial areas can readily be
more than two orders of magnitude above those at
rural/agricultural locations. Whether these risks are
actually realized is entirely dependent on the
magnitude and frequency of exposure. With the
exception of particular situations, residential expo-
sures to contaminated soils at properly managed
hazardous waste sites seem unlikely. The aforemen-
tioned daily soil ingestion rates routinely employed
for risk assessment are only 100 and 200 mg for adults
and children respectively, and even at the RME
(reasonable maximum exposure), only a small fraction
of this daily value will include fugitive dusts and
tracked soil from the contaminated area. Exceptions
would include industrial workers at the site and
residents that are not aware of the hazard and
frequently enter the contaminated area. This can be
particularly serious for children who venture on to the
site for recreational purposes. The scenario of the ill-
informed resident from the nearby community can be
a serious issue, and proper risk communication can be
difficult. For example, cancer risk communication in
Canada’s arctic, a region that contains orphaned
military sites contaminated with PCBs and petroleum
hydrocarbons, requires the creation of new words in
the local language to explain terms like ‘‘pollution’’
and ‘‘contamination’’.
An additional 23% of the studies examined were
motivated by genotoxic hazard assessment and
remediation evaluations, and 5% investigated the
likelihood of future hazards associated with current or
planned waste disposal practices. For example, four
studies examined the genotoxic hazards associated
with long-term applicatsions of municipal waste-
waters or sewage sludge on agricultural lands
[60,165,176,179]. The summarized remediation
efforts (Table 21) suggest that some bioremediation
strategies employed for soils contaminated with
recalcitrant genotoxins (e.g., nitroaromatic explosives,
creosote) are unable to remove the hazard. Moreover,
in some cases hazard was notably increased. Although
the mechanistic details controlling this phenomenon
are not known, it is reasonable to assume that the
phenomenon results from the formation of transient
breakdown products that are, in toto, more mutagenic
than the starting material. The supposition is
supported by the detailed analytical works of
Lundstedt et al. and Brooks et al. [22,353]. Their
analyses of bioslurry treatments for gas-works soil and
soil from a wood-preservation site, respectively,
showed the formation of PAH derivatives, some of
which are known to be mutagenic (e.g., ketones and
quinones). Although remediation of soils contami-
nated with other PAH-rich wastes can induce
relatively short-lived increases in mutagenic hazard
(i.e., weeks to months), the hazards appear to be
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 289
effectively removed within one year. Thus, if the site is
properly managed (i.e., restricted access, runoff
collection, HDPE liner), and site recovery is not
urgent, it would appear that the less sophisticated
remediation options (e.g., land treatment, biopile,
compost) are the most parsimonious. Nevertheless, it
should be noted that many researchers advocating the
use of bioreactor treatments such as the bioslurry,
emphasize the advantage of short treatment time
[346]. It should also be noted that this type of
strategy would not be appropriate for sites that are
contaminated with mutagenic metals. Changes in
oxidation state, speciation and complexation could
liberate mutagenic metals, and effective remediation
strategies should include a technique such cation
solidification to immobilize mutagenic metals [337].
A small number of studies specifically evaluated
protocols for the extraction and handling of con-
taminated soils for mutagenicity assessment. For
example, Wang et al. compared four solvent systems
(i.e., hexane/acetone, DCM/acetone, hexane/2-propa-
nol, DCM) for their ability to extract S9-activated
Salmonella mutagens (TA98 and TA100) from soils
collected at a hazardous landfill [423]. The results
showed a relatively low response on both strains for
the DCM extracts, and roughly equivalent responses
for the other three solvents on TA100. The TA98
results indicated that hexane/acetone and DCM/
acetone were more effective than hexane/2-propanol.
The results summarized in Table 7 indicate that DCM
frequently provides samples that are more mutagenic
on TA98 (with S9). Thus, it is not possible to select a
solvent or solvent system that is suitable for all
situations. Similar conclusions have been recorded for
other complex environmental matrices. Lee et al.
found that acetone was superior to DCM for extracting
Salmonella mutagens from airborne particulate
matter; however, Kiel et al. did not detect any
difference in the Salmonella mutagenicity of coal-tar
contaminated sediment extracts prepared using
DCM or acetone/hexane [424,425]. Solvent choice,
therefore, must be study-specific and consider the
source of the soil sample and the physical-chemical
properties of the purported contaminants. However, it
should be noted that the results of several works such
as that of Ehrlichmann et al. [174] on the bacterial
genotoxicity of aqueous soil extracts, and many
studies that employed plant assays, definitively
indicate that solvent extraction is essential for reliable
genotoxicity assessment of soils contaminated with
organic contaminants.
In the absence of a generally applicable rule
regarding solvent choice, it seems prudent to choose a
solvent or system that can dissolve a wide range on
compounds (e.g., polar aromatics, non-polar aro-
matics, neutral compounds, etc.). A recent investiga-
tion by Lundstedt et al revealed that newer methods
such as accelerated solvent extraction (ASE) with
binary solvent mixtures such as acetone/hexane (1:1,
v/v) are preferred for effective extraction of organics
from PAH-contaminated soils [426]. Moreover, these
authors noted that the ASE–hexane/acetone combina-
tion reduces solvent use, avoids chlorinated solvents,
and provides an extract that is easily handled in
subsequent clean-up or fractionation steps.
Although the data discussed in this review indicate
that the Salmonella mutagenicity test is the most
commonly employed bioassay for the examination of
contaminated soils (Fig. 1), few researchers have
actually attempted comparative evaluations of assay
performance. Three studies evaluated the efficacy of
the Salmonella assay for assessments of soils
contaminated with chlorinated substances. Cizmas
et al. noted that the prophage induction assay detected
significant genotoxic activity in an acid fraction of a
wood preserving waste extract (PCP �472 ppm) that
did not induce a positive response on TA98 with S9
[151]. The examination of a PCB-contaminated soil
also showed differential responses between the
Salmonella mutagenicity assay and the prophage
induction test, and the authors attribute the difference
to the previously observed sensitivity of the prophage
induction assay to chlorinated compounds [41].
Donnelly et al. compared the rec� differential survival
assay and the Salmonella assay (TA98 with S9) for
examinations of a wood preserving waste extract and
revealed strong responses for both assays to acid and
base fractions [146]. However, although the neutral
fraction induced the strongest response on TA98 with
S9, no significant response was obtained for the rec�
assay. The same waste extract was employed to
compare the Salmonella assay with mutagenicity and
clastogenicity endpoints in haploid and diploid
Aspergillus sp. [317]. The results revealed potent
mutagenicity in haploid Aspergillus (with S9) for
the basic fraction, and potent responses in the
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345290
diploid assay to both the acid and base fractions. This
response pattern is quite distinct from the aforemen-
tioned pattern observed in Salmonella TA98 (i.e.,
maximum for the neutral fraction). Collectively, these
results indicate that complex samples such as extracts
of contaminated soil will yield different responses in
different assay systems with different endpoints. This
is not surprising since wood preserving waste is known
to contain a complex mixture of chlorinated com-
pounds (e.g., PCP), PAHs, PAH derivatives, and
heterocyclic aromatic compounds [427]. A compara-
tive evaluation of TA98 mutagenicity and differential
survival in repair deficient E. coli by Aleem and Malik
[60] indicated very similar responses for a series of
extracts from soils irrigated with industrial and
municipal wastewaters.
An interesting study by Malachova et al. suggested
that Salmonella TA98 (with and without S9) is more
sensitive than the SOS Chromotest response, and the
differential responses could yield completely opposite
conclusions regarding the efficacy of a bioremediation
exercise for a coke oven contaminated soil [153].
Although it is not clear whether these differential
responses in genotoxicity are generally applicable, it is
clear that the conclusions of the research can be
seriously effected by bioassay choice. However, it
should be noted that a detailed overview on the
mutation spectra of complex environmental mixtures
(e.g., drinking water, urban air, cigarette smoke)
suggests that observed differences between the
response patterns of assays with similar endpoints
(i.e., mutation) will be overshadowed by similarities in
the induced mutation pattern [129].
The collected data also indicate that collectively,
plant assays are as popular as the Salmonella
mutagenicity test for assessing the genotoxic hazards
of contaminated soils. The endpoints routinely
employed include gene mutation in stamen hairs
(Tradescantia) or second-generation embryos (Arabi-
dopsis), chromosome aberrations in root tips (Allium),
MN in Tradescantia tetrads, and sister chromatid
exchanges (Vicia). The method of exposure used in the
plant publications is highly varied and includes
exposures to aqueous extracts or leachates, soil
slurries, organic extracts, as well as direct soil contact.
Some researchers performed comparative evalua-
tions of the popular plant assay systems. Knasmuller et
al. noted that the Vicia MN assay was incapable of
detecting genotoxicity in metal contaminated soils
that was readily detected using the Tradescantia MN
assay (direct contact) [76]. That study, and the results
of the data analyses conducted here, also noted
relatively small differences in the mean Tradescantia
MN response to aqueous soil leachates. As already
mentioned (section 8.6), the study by Cotelle et al
noted that comparisons of MN assays in Vicia, Allium
and Tradescantia revealed both equivalent responses
(i.e., PAH contamination) and differential responses
(i.e., Vicia > Allium > Tradescantia for PCB contam-
ination) [83]. Three studies by Cabrera et al. compared
the Allium aberration test, the Tradescantia SHM test
and the Tradescantia MN test for examinations of
leachates from samples of composted garbage, soils
irrigated with wastewater, and soil from a municipal
landfill [66,67,283]. The results indicated clear
differences in the response pattern, with a stronger
response for the Tradescantia MN assay, and weaker
or insignificant responses for the Tradescantia SHM
and Allium assays. Kong and Ma’s examination of
aqueous soil extracts for sites contaminated with
pesticides or metals revealed an identical response
patter for the Allium CA assay and the Tradescantia
MN assay [247]. Comparisons of the Tradescantia
SHM assay and the waxy mutation assay in Zea mays
for analysis of soils in the vicinity of a lead smelter, a
petrochemical complex, and an oil refinery (direct-
contact exposure) revealed very similar response
patterns, but increased sensitivity and a far greater
response range for the Zea mays assay [70,71].
Although it is difficult to draw general conclusions
from these comparative assessments, the results
presented in this review do permit some general
comments. First, it appears that the direct-contact
assessment is far more effective for soil genotoxicity
assessment than immersion of cuttings or roots in
aqueous leachates or extracts. Unfortunately, there
was not enough data for a definitive evaluation of
organic extraction, although some limited evidence
(e.g., [66,68,69]) suggests that organic extracts may be
suitable for the analysis of soils contaminated with
combustion by-products (e.g., PAHs) or other organ-
ics. Second, the Allium CA assay and Zea mays waxy
locus assay are preferable to the other assays (i.e.,
Tradescantia MN, Tradescantia SHM). The mean
response ranges of these assays are approximately 5-
to 10-fold above control, with values reported in the
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 291
literature as high as 25-fold above control [71]. More-
over, average cross-study variations in mean responses
for these assays are relatively low. Despite these advan-
tages, the Allium CA assay and the waxy mutation
assay in Zea are more technically challenging. Scoring
of CAs requires specialized training, and cultivation of
Zea mays requires large amounts of space and 6–12
weeks before the results can be obtained (i.e., seed to
tassel) [71]. However, it might be noted that the
extended direct contact exposure employed in the
Zea mays waxy locus assay may be partly responsible
for the responsiveness of this assay system.
Very few researchers have attempted to evaluate the
popular Salmonella endpoint via comparisons with
mammalian in vitro or in vivo endpoints. Berthe-Corti
et al. observed close correspondence between Salmo-
nella mutagenicity (TA98 and TA100 with and
without S9) and the induction of hprt mutations in
Chinese Hamster V79 cells for a DMSO/EtOH extract
of a munitions contaminated soil [65], and additional
chemical analysis showed the presence of TNT,
hexogen and several DNTs. Wesp et al. examined
fractions of soils contaminated with emissions from
gasoline- and diesel-powered vehicles and showed
significant induction of SCEs in cultured human
lymphocytes (with and without S9), but no induction
of MN in mouse bone marrow following gavage
exposure [116]. Simultaneous analyses showed potent
direct-acting Salmonella TA98 activity attributable to
nitroarenes, and the authors reconcile the lack of
correspondence using suppositions about threshold
concentrations and antagonism of mixtures compo-
nents. However, the observed differences between the
microbial and mammalian results are likely caused by
differences in the ability to reduce the nitroaromatics.
Nitroreduction is a critical step in the activation of
nitroarenes and the specific activity in bacterial
systems is known to be far greater than that of
mammalian systems [428–430]. Therefore, confirma-
tion of animal hazard via in vivo testing appears to be
an essential step in hazard confirmation and risk
assessment. Surprisingly, only one published study
successfully demonstrated an adverse effect following
ingestion of contaminated soil particles. Dietary
exposures of rats to coal-tar amended soils showed
significant induction of bulky (i.e., PAH) DNA
adducts in both liver and lung [326], thus confirming
genotoxic hazard.
A surprisingly small proportion of the reviewed
studies (�8%) were primarily concerned with identify-
ing the major source(s) of the soil genotoxicity. Several
Japanese studies investigated the mutagenic hazards of
urban soils and evaluated the contributions of DNPs
from diesel emissions, as well as PAHs from
automobile exhaust and studded tires [50,51,119,
124,431]. These works ultimately showed that DNPs
can account for up to 27% of the direct acting TA98
mutagenic activity in extracts of urban soils [50–
52,124]. Using the combined data from five studies, the
analysis conducted here (Fig. 5) confirmed a strong
empirical relationship between direct-acting TA98
mutagenicity and DNP contamination across 37 sites.
Two German studies investigated the sources of
mutagenic activity in a variety of soils and concluded
that urban soils are contaminated via airborne deposi-
tion of combustion by-products such as PAHs and
related compounds (e.g., 2-nitrofluorene) [55,116].
This conclusion is similar to that of Schoen et al. [432]
who concluded that urban soils collected from the St.
Lawrence River riparian region downstream from
Montreal are primarily contaminated with unsubsti-
tuted, homocyclic PAHs. Although this study did not
include chemical analysis, TA98 and TA100 S9-
activated Salmonella mutagenicity were roughly
equivalent and the metabolically enhanced strains
YG1041 and YG1042 showed no increase in response.
It is interesting to note that although one of the
aforementioned German studies [116] highlighted
combustion by-products from gasoline and diesel
powered vehicles as the likely source of the soil
mutagenicity, they did not detect a significant correla-
tion between soil mutagenic activity (TA98 with S9)
and PAH concentration. Moreover, the authors con-
cluded that the bulk of the mutagenic activity (up to
54%) in fractions of soils exposed to motorway exhaust
is associated with polar organics such as nitroarenes.
Examinations of coal-tar amended soils by
Alexander et al. also failed to reveal a relationship
between mutagenic activity and the concentration of
selected carcinogenic PAHs [90]. The results pre-
sented in this review (Fig. 4) did show a statistically
significant (p < 0.0001), albeit fairly weak, relation-
ship (r2 = 0.17 to 0.25) between S9-activated TA98
mutagenicity and soil PAH concentration (59–80
observations from 13 studies). Close scrutiny of the
relationship suggests that restricted subsets of the data
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345292
are unlikely to yield a statistically significant relation-
ship. Therefore, although 17–25% of the cross-study
variations in S9-activated frameshift activity can be
explained by PAH concentration, the results of
individual studies indicate that the commonly mea-
sured mutagenic PAHs (e.g., benzo[a]pyrene, ben-
zo[b]fluoranthene, dibenz[a,h]anthracene) cannot
account for the bulk of soil TA98 S9-activated
mutagenic hazard.
The results presented here also showed empirical
relationships between mutagenic and clastogenic
effects in plants (e.g., Allium and Arabidopsis) and
radionuclide or chemical contamination (e.g., 137Cs or
PAHs). These relationships confirm the use of plants
for monitoring sites contaminated by radioactive
substances. Although the relationship between Tra-
descantia MN and PAH contamination would not be
significant without the Baud-Grasset et al. [120]
observation, it is intriguing that Tradescantia appears
to have the metabolic capacity to activate the
mutagenic components in creosote-contaminated soil.
Numerous studies of creosote-contaminated soils have
noted potent Salmonella mutagenicity (TA98 and
TA100) that is greatly enhanced by the addition of S9,
and further enhanced by S9 concentrations above the
standard concentration (i.e., 30% S9 v/v in mix)
[42,146]. Several researchers have documented the
ability of plants and algae to metabolize and activate
mutagenic substances including PAHs [433–436], and
the observed plant responses summarized throughout
this work indicates that selected plants (e.g., Zea
mays) can be useful for assessing the likelihood of
adverse ecological effects caused by DNA damage.
A surprisingly small number of the reviewed
studies employed bioassay-directed fraction to isolate
and identify the putative mutagens in contaminated
soils. The strategy has been successfully employed to
isolate novel mutagens in contaminated sediments and
surface waters [23,144,437]; however, although many
studies employed a coarse fractionation procedure
(i.e., acid, base, neutral) in their analysis of soil
extracts [42,118,146,151,317], very few researchers
have attempted detailed bioassay-directed fractiona-
tion and compound identification. This may be due to
the complexity of contaminated soils and the
analytical challenge of isolating and identifying soil
mutagens. Two studies that used a thorough bioas-
say-directed fractionation approach to isolate and
identify Salmonella mutagens in soils contaminated by
wood-preserving wastes (e.g., creosote) were unable to
definitively identify the putative mutagens. Brooks et al.
subjected 40 fractions of untreated and bioremediated
creosote-contaminated soil to detailed mutagenicity
and chemical analyses, and the final results revealed
over 100 compounds or compound groups several of
which have never been assessed for mutagenic activity
[22]. Nevertheless, the results did identify a range of
known Salmonella mutagens. These included PAHs, N-
containing heterocyclics, S-containing heterocyclics,
and oxygenated PAH derivatives (e.g., quinones)
[134,141,142,319,376,378,379]. Wesp et al. employed
a less sophisticated fractionation procedure and a
variety of metabolically manipulated Salmonella
strains (e.g., TA98NR, YG1021, YG1024, YG1026
and YG1029) to determine the chemical class of
mutagens in soils exposed to automobile emissions
[116]. The results showed that most of the mutagenic
activity (55–65%) is attributable to polar aromatics
such as nitroarenes, and only 10% is attributable to non-
polar neutrals such as homocyclic, unsubstituted PAHs.
Therefore, although few studies have employed
detailed fractionation protocols to isolate and identify
soil mutagens, these studies did confirm that a
substantial fraction of the mutagenic activity in urban
soils is attributable to nitroarenes and other PAH
derivatives.
It is unfortunate that few studies even attempt to
interpret genotoxicity results in a broader, system-
level focus. For example, an examination of the soil
depth where mutagenic hazard is highest would permit
researchers investigating urban sites to determine
whether contamination arriving via dry deposition is
recent. One study did present a depth profile of
Salmonella mutagenic activity for soils exposed to
vehicular emissions [116]. The results reveal a clear
decline in mutagenic activity (with and without S9,
several strains) for depths below 10 cm, and some
indication of a maximum between 5 and 10 cm.
Although this likely indicates maximal deposition of
airborne mutagens at some point in the past, it is
impossible to assess the time frame without detailed
information about the site’s physical history. At
industrial or hazardous waste dumpsites, depth
profiles can determine the ability of the contamination
to move through the various soil horizons eventually
impacting the groundwater. Examinations of soil
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345 293
borings collected at an abandoned chemical manu-
facturing site by Donnelly et al. revealed slightly
higher S9-activated TA98 mutagenicity in surface
soils (0–0.6 m), as compared to samples collected
between 1 and 4 m [47]. However, the samples
collected between 1 and 4 m also showed substantial
mutagenic activity, and the authors were unable to
chemically identify the mutagenic substances or offer
an explanation for the noted activity profile.
Although other variables such as soil texture, grain
size distribution and organic matter content would be
expected to effect extraction efficiency and the
bioavailability of mutagenic metals, few studies
attempted to assess the impact of soil properties on
the results obtained. Some studies do note sampling
depth [50,54,60,68,77,114,115,118,193], and/or soil
texture [40,48] and grain size distribution
[40,48,50,65]; however, this information is rarely used
to interpret the results. One exception is the study of
Majer et al. that investigated correlations between soil
mutagenicity (MN induction in Tradescantia by direct-
contact) and a variety of parameters used to describe the
vitality of the soil microbial community [77]. The
results showed negative correlations between MN
induction and both microbial biomass and dehydro-
genase activity, and a positive correlation between MN
induction and metabolic quotient (i.e., microbial
activity to biomass ratio). This may simply indicate
that the mutagenic metals in the samples are also toxic
to bacteria, accounting for a drop in overall biomass and
enzyme activity, and an increase in overall metabolic
quotient when MN induction activity is observed.
Additional analysis showed a positive correlation
between metabolic quotient and As concentration.
None of the papers examined attempted to interpret
the results obtained in an ecosystem health or
ecotoxicological context. One would suspect that an
enhancement in the mutagenic hazards of a terrestrial
environment would have repercussions for the
indigenous biota. Mutation frequencies should
increase and this would be expected to have
evolutionary as well as ecological consequences.
Several researchers have suggested that continued
accumulation of deleterious mutations in sufficiently
small populations can initiate a phenomenon, known
as mutational meltdown, that has the ability to drive
populations to extinction [438–440]. The likelihood of
this phenomenon at sites heavily contaminated with
mutagenic substances is completely unknown. How-
ever, it is known that contaminated soils can induce
DNA damage and CAs in indigenous rodents
[326,441], DNA damage in earthworms [442], and
changes in microbial community structure [443].
Moreover, recent finding show that chronic in situ
exposures to radionuclide or chemical contamination
is capable of inducing epigenetic and heritable
adaptive responses in vertebrates (e.g., fish) and
plants (i.e., Arabidopsis) [444,445].
12. Conclusions and future prospects
To our knowledge, the first published work
documenting the mutagenic hazards of soils was
published in 1981 [446], less than one year after the
US government enacted CERCLA, also known as the
Superfund Act. This act provided the motivation and
funding for research investigating the mutagenic
hazards associated with contaminated land. Since
1981, some 118 publications encompassing over 1300
assessments of soil genotoxicity have been published.
Most of these studies were motivated by a need for
effective assessment of genotoxic hazard. However, an
assessment of the actual risk for mutagenic or
carcinogenic effects associated with contaminated
soils requires accurate knowledge of exposure.
Although current risk assessment guidelines do
provide soil exposure values for adults and children
in different settings, it is extremely difficult to assess
the proportion of a daily exposure that would be
represented by soil particles from a contaminated area.
Nevertheless, the precautionary approach seeks to
eliminate all exposures and liability for damages, and
this elimination can be achieved via access restriction,
site remediation and rehabilitation.
In urban settings widespread exposure is far more
likely and accurate assessment of risk requires more
reliable exposure estimates for both adults and
children in a variety of settings (i.e., residential,
commercial, institutional, outdoor). Although values
are available in the literature, there are technical
challenges associated with values that differentiate
between exposures to soil particles and exposures to
indoor dust [415,447] (see also Maertens et al., this
issue [448]). Very little research has attempted to
investigate the effects of contaminated urban soils
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345294
following non-dietary ingestion or inhalation. The
only available studies suggest a possibility of
greatly enhanced levels of DNA damage in liver
and lung resulting from ingestion of PAH-contami-
nated material [326]. Thus, efforts to improve the
accuracy of mutagenicity and carcinogenicity risk
estimates for urban residents exposed to soils
contaminated through dry deposition of airborne
particulate matter seems to be a promising area for
further research.
Although a great deal of progress has been made
regarding the ability to remediate and rehabilitate sites
contaminated with mutagenic substances such as
munitions, creosote and coal tar, it is not clear whether
chemical-specific monitoring of remediation progress
is effective. In some cases, chemical monitoring
showed dramatic declines in prioritized substances
(e.g., TNT), although total genotoxic hazard remained
largely unchanged. Perhaps future research should
employ bioassays to monitor remediation progress,
and subsequently employ a bioassay-directed fractio-
nation strategy to isolate and identify putative
mutagens that are problematic. These hitherto
unknown compounds can be prioritized for subsequent
remedial action and inclusion in risk assessments. In
addition, modern genetic tools could be used to
identify the genetic and biochemical attributes of a
remediation system that enhance the likelihood of risk
removal. For example, biochemical screening of
environmental isolates has permitted the isolation
and identification of pentaerythritol tetranitrate
(PETN) and glycerol trinitrate, two bacterial nitrate
ester reductases that are capable of degrading
mutagenic explosives [449]. These two enzymes have
recently been integrated into the genome of tobacco
and this transgenic plant can provide an effective and
affordable means to remediate soils contaminated with
explosives [450]. Recent work by He et al. has
employed 16S rRNA gene-based tools to identify a
population of Dehalococcoides capable of completely
mineralizing chlorinated substances such as tetra-
chloroethene and vinyl chloride [360,451]. The
organism is now commercially available under the
name Bio-Dechlor INOCULUMTM, a lyophilized
microbial consortium that can accelerate mineraliza-
tion of chlorinated solvents in contaminated soils (see
www.regenesis.com).
Finally, it is clear that there is a paucity of
information on the ecological hazards of genotoxic
soils. Although a few studies have documented effects
on earthworms and rodents [441,442], the nature and
magnitude of the effects on community structure and
population viability are completely unknown. Assess-
ment of exposure levels, bioavailability, and effects on
terrestrial organisms is another promising area for
future research.
Acknowledgements
We would like to thank several people for providing
essential assistance. Prof. Kirby Donnelly of Texas
A&M University, Prof. Katerina Malachova of the
University of Ostrava, and Dr. Osamu Endo of the
Japanese National Institute of Public Health provided
access to unpublished information. Dr. Tamara
Grummt of the German Umweltbundesamt,
Außenstelle Bad Elster provided information about
contaminated sites in Germany. Prof. Takeshi Ohe of
Kyoto Womens University provided several Japanese
publications. The Government of Canada translation
office provided translation of Spanish, Italian and
Russian manuscripts. The US National Research
Council (under the Research Associateship Program),
the Natural Sciences and Engineering Research
Council of Canada (under the visiting fellowship
program), and Health Canada (under the Investing in
Biotechnology program) provided funding. This work
was also partly funded by the US Environmental
Protection Agency. It has been subjected to review by
the National Health and Environmental Effects
Research Laboratory and approved for publication.
Approval does not signify that the contents reflect the
views of the Agency, nor does mention of trade names
or commercial products constitute endorsement or
recommendation for use.
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Appendix A
Salmonella mutagenicity data collected from the literature
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g dry
soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
I. Sites not contaminated by industrial/urban activities (i.e., rural and/or agricultural)
Row crops (hops) 504.0 504.0 636.0 986.0 Ac/Hex NAd PAHse/pesticidesf PAHs: 2.5 [57]g
Row crops (asparagus) 493.0 471.0 526.0 756.0 Ac/Hex NA PAHs/pesticides PAHs: >0.1 [57]
Row crops (rye) 132.0 137.0 230.0 241.0 Ac/Hex NA Pesticides [57]
Row crops (maize) 208.0 214.0 285.0 241.0 Ac/Hex NA PAHs/pesticides PAHs: >0.07 [57]
Row crops (oats) 99.0 93.0 153.0 186.0 Ac/Hex NA [57]
Pasture 132.0 137.0 252.0 263.0 Ac/Hex NA PAHs PAHs: >0.01 [57]
Meadow 115.0 115.0 219.0 351.0 Ac/Hex NA [57]
Rincon series loam 36.0 298.0 33.0 73.0 ACN NA [58]
Rincon series loam NA 72.0 NA 22.0 MetOH NA [58]
Rincon series loam NA 33.0 NA 34.0 DCM NA [58]
Rincon series loam NA 4.0 NA 9.0 Water (pH = 2) NA [58]
Field (no agriculture) NA 254.0 NA 66.0 ACN NA [58]
Field (20 cm depth) NA 119.0 NA 49.0 ACN NA [58]
Field 2 (no agriculture) NA 232.0 NA 46.0 ACN NA [58]
Garden 1 (organic) NA 141.0 NA 83.0 ACN NA [58]
Garden 2 (organic) NA 217.0 NA 44.0 ACN NA [58]
Forest edge NDh 50.0 ND 230.0 DCM NA [432]
Bastrop soil (rangeland) 88.7 98.8 69.9 132.9 DCM 0.23 Pesticides [40,48,49]
Sassafrass soil (row crops) 1.0 4.0 6.3 3.9 DCM 0.025 Pesticides [40,49]
Norwood soil (row crops) 0.3 2.0 1.9 0.8 DCM 0.057 Pesticides [40,42,49,166]
Rural forest (Wales) NTi 78.0 NT 95.0 DCM NA PAHs PAHs: 2.2 [39]
Pasture (Wales) NT 46.0 NT 106.0 DCM NA PAHs PAHs: 0.2 [39]
Grazing land (Wales) NT 42.0 NT 96.0 DCM NA PAHs PAHs: 0.3 [39]
Ploughed/grazing (Wales) NT 39.0 NT 93.0 DCM NA PAHs PAHs: 0.3 [39]
Weswood soil (row crops) 42.0 14.0 NA NA MetOH 1.31 [64,165]
Weswood soil (row crops) 2.0 2.0 NA NA DCM 0.16 [64,165]
Weswood soil (row crops) 0.3 2.0 NA NA DCM 0.057 [48]
Weswood soil (row crops) 19.0 3.0 NT NT DCM/MetOH 1.48 [64]
Parkland (Azerbaijan 11) NT 7.0 NT NT Ac/Hex 9.39 PAHs PAHs: 14.5 [56]
Padina soil (pasture) 12.0 2.0 NA NA MetOH 0.46 [165]
Padina soil (pasture) 7.0 6.0 NA NA DCM 0.57 [165]
Hechtsheim 155.0 220.0 340.0 420.0 ACN NA [55]
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
29
6Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
Hechtsheim 145.0 142.0 205.0 340.0 MetOH NA [55]
Hechtsheim 110.0 205.0 545.0 1040.0 DCM NA [55]
Hechtsheim 125.0 260.0 335.0 435.0 Hex/Ac NA [55]
Hechtsheim 54.0 120.0 100.0 174.0 Ac/Tol/
MetOH
NA [55]
Laubenheim 25.0 40.0 ND ND ACN NA [55]
Laubenheim 28.0 53.0 ND 22.0 DCM NA [55]
Laubenheim 40.0 40.0 ND 38.0 Hex/Ac NA [55]
Ober-Olm 35.0 80.0 ND 55.0 ACN NA [55]
Ober-Olm 37.0 73.0 107.0 21.0 DCM NA [55]
Ober-Olm 30.0 140.0 140.0 ND Hex/Ac NA [55]
Alzey (cereals) 21.8 ND ND ND Hex/Ac NA [55]
Bubenheim (cereals) 405.0 ND ND ND Hex/Ac NA [55]
Bubenheim (meadow) ND 30.0 ND ND Hex/Ac NA [55]
Bubenheim (viniculture) ND 765.8 ND 102.5 Hex/Ac NA [55]
Bindlach (cereals) 40.2 23.8 ND ND Hex/Ac NA [55]
Bindlach (meadow) 430.0 12.4 ND ND Hex/Ac NA [55]
Essenheim I (cereals) 61.0 22.7 ND ND Hex/Ac NA [55]
Essenheim I (viniculture) 45.0 45.4 ND ND Hex/Ac NA [55]
Essenheim II (orchard) 9.2 52.0 ND ND Hex/Ac NA [55]
Essenheim II (cereals) 60.5 86.4 ND ND Hex/Ac NA [55]
Finthen, MZ (cereals) 5.0 67.0 ND ND Hex/Ac NA [55]
Finthen, MZ (orchard) 44.9 102.2 ND ND Hex/Ac NA [55]
Gaualgesh, NR (forest) 116.6 226.9 ND 261.5 Hex/Ac NA [55]
Gaualgesh, NR (cereals) 18.0 13.9 ND ND Hex/Ac NA [55]
Hechtsheim, MZ ND 117.8 ND ND Hex/Ac NA [55]
Ingelheim (orchard) 65.7 73.3 ND 78.1 Hex/Ac NA [55]
Ingelheim (orchard) 155.0 73.0 ND ND Hex/Ac NA [55]
Laubenheim I, MZ (cereals) 79.6 75.0 ND ND Hex/Ac NA [55]
Laubenheim I, MZ (fallow) ND ND 475.0 115.0 Hex/Ac NA [55]
Laubenheim I, MZ (meadow) ND 135.6 ND ND Hex/Ac NA [55]
Laubenheim II (cereals) 42.8 169.8 ND 195.0 Hex/Ac NA [55]
Laubenheim II (cereals) ND 300.0 ND ND Hex/Ac NA [55]
Laubenheim II (forest) ND 60.2 ND ND Hex/Ac NA [55]
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
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67
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04
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27
–3
45
29
7
Mainzer sand, NR (forest) 28.2 31.0 ND 140.0 Hex/Ac NA [55]
Mainzer sand, NR (fallow) 100.0 160.2 ND ND Hex/Ac NA [55]
Neukirchen (meadow) 30.0 ND ND ND Hex/Ac NA [55]
Ober-Olm (orchard) ND ND 265.0 ND Hex/Ac NA [55]
Ober-Olm (forest) 266.9 60.8 263.5 ND Hex/Ac NA [55]
Ober-Olm (cereals) ND 82.8 ND ND Hex/Ac NA [55]
Rauhenberg (cereals) 44.6 ND ND ND Hex/Ac NA [55]
Scheckenhof (cereals) 480.0 86.4 ND ND Hex/Ac NA [55]
Scheckenhof (meadow) 144.0 181.6 ND ND Hex/Ac NA [55]
Bubenheim (rye) 43.0 12.0 33.0 25.0 Hex/Ac NA [55]
Bubenheim (rye) 50.0 51.0 52.0 54.0 Hex/Ac NA [55]
Bubenheim (rye) 117.0 70.0 85.0 70.0 Hex/Ac NA [55]
Bubenheim (rye) 418.0 71.0 146.0 145.0 Hex/Ac NA [55]
Bubenheim (rye) 80.0 99.0 75.0 121.0 Hex/Ac NA [55]
Bubenheim (rye) 43.0 35.0 70.0 91.0 Hex/Ac NA [55]
Bubenheim (rye) 165.0 125.0 270.0 80.0 Hex/Ac NA [55]
Essenheim I (rye) 7.0 18.0 79.0 36.0 Hex/Ac NA [55]
Essenheim I (rye) 105.0 44.0 115.0 105.0 Hex/Ac NA [55]
Essenheim I (rye) 75.0 75.0 185.0 130.0 Hex/Ac NA [55]
Essenheim I (rye) 245.0 130.0 249.0 108.0 Hex/Ac NA [55]
Essenheim I (rye) 85.0 55.0 58.0 60.0 Hex/Ac NA [55]
Essenheim I (rye) 100.0 33.0 80.0 95.0 Hex/Ac NA [55]
Essenheim I (rye) 130.0 60.0 144.0 315.0 Hex/Ac NA [55]
Essenheim II (rye) 31.0 30.0 135.0 123.0 Hex/Ac NA [55]
Essenheim II (rye) 52.0 51.0 150.0 94.0 Hex/Ac NA [55]
Essenheim II (rye) 255.0 70.0 242.0 86.0 Hex/Ac NA [55]
Essenheim II (rye) 185.0 216.0 245.0 167.0 Hex/Ac NA [55]
Essenheim II (rye) 91.0 135.0 60.0 65.0 Hex/Ac NA [55]
Essenheim II (rye) 76.0 30.0 95.0 40.0 Hex/Ac NA [55]
Essenheim II (rye) 175.0 158.0 185.0 235.0 Hex/Ac NA [55]
Finthen, MZ (rye) 42.0 19.0 90.0 31.0 Hex/Ac NA [55]
Finthen, MZ (rye) 30.0 40.0 31.0 59.0 Hex/Ac NA [55]
Finthen, MZ (rye) 52.0 54.0 175.0 230.0 Hex/Ac NA [55]
Finthen, MZ (rye) 145.0 119.0 107.0 151.0 Hex/Ac NA [55]
Finthen, MZ (rye) 116.0 107.0 156.0 178.0 Hex/Ac NA [55]
Finthen, MZ (rye) 89.0 102.0 48.0 95.0 Hex/Ac NA [55]
Finthen, MZ (rye) 54.0 85.0 95.0 169.0 Hex/Ac NA [55]
Gaualgesheimer (rye) 12.0 18.0 55.0 66.0 Hex/Ac NA [55]
Gaualgesheimer (rye) 49.0 36.0 88.0 44.0 Hex/Ac NA [55]
Gaualgesheimer (rye) 19.0 42.0 55.0 65.0 Hex/Ac NA [55]
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
29
8
Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
N ppm dry weightc
Gaualgesheimer (rye) 120.0 191.0 163.0 98.0 Hex/Ac NA [55]
Gaualgesheimer (rye) 64.0 110.0 135.0 81.0 Hex/Ac NA [55]
Gaualgesheimer (rye) 23.0 30.0 45.0 33.0 Hex/Ac NA [55]
Gaualgesheimer (rye) 70.0 50.0 155.0 175.0 Hex/Ac NA [55]
Hechtsheim, MZ (rye) 22.0 17.0 60.0 87.0 Hex/Ac NA [55]
Hechtsheim, MZ (rye) 73.0 155.0 235.0 110.0 Hex/Ac NA [55]
Hechtsheim, MZ (rye) 90.0 96.0 227.0 124.0 Hex/Ac NA [55]
Hechtsheim, MZ (rye) 95.0 38.0 88.0 205.0 Hex/Ac NA [55]
Hechtsheim, MZ (rye) 100.0 152.0 78.0 197.0 Hex/Ac NA [55]
Hechtsheim, MZ (rye) 37.0 105.0 56.0 170.0 Hex/Ac NA [55]
Hechtsheim, MZ (rye) 110.0 135.0 90.0 355.0 Hex/Ac NA [55]
Laubenheim I, MZ (rye) 48.0 12.0 145.0 19.0 Hex/Ac NA [55]
Laubenheim I, MZ (rye) 50.0 18.0 59.0 135.0 Hex/Ac NA [55]
Laubenheim I, MZ (rye) 55.0 125.0 310.0 85.0 Hex/Ac NA [55]
Laubenheim I, MZ (rye) 70.0 255.0 130.0 60.0 Hex/Ac NA [55]
Laubenheim I, MZ (rye) 40.0 570.0 530.0 140.0 Hex/Ac NA [55]
Laubenheim I, MZ (rye) 41.0 77.0 200.0 121.0 Hex/Ac NA [55]
Laubenheim I, MZ (rye) 75.0 155.0 265.0 105.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 32.0 26.0 108.0 31.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 85.0 98.0 73.0 132.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 60.0 79.0 136.0 154.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 120.0 120.0 207.0 291.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 97.0 194.0 135.0 220.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 93.0 58.0 140.0 73.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 55.0 105.0 270.0 135.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 32.0 10.0 85.0 15.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 60.0 47.0 153.0 124.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 130.0 15.0 207.0 79.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 125.0 72.0 370.0 325.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 45.0 125.0 595.0 88.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 14.0 51.0 70.0 28.0 Hex/Ac NA [55]
Laubenheim II, MZ (rye) 70.0 65.0 200.0 110.0 Hex/Ac NA [55]
ame
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
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67
(20
04
)2
27
–3
45
29
9
Ober-Olm (rye) 31.0 16.0 89.0 88.0 Hex/Ac NA [55]
Ober-Olm (rye) 40.0 61.0 78.0 140.0 Hex/Ac NA [55]
Ober-Olm (rye) 25.0 42.0 155.0 210.0 Hex/Ac NA [55]
Ober-Olm (rye) 90.0 156.0 262.0 310.0 Hex/Ac NA [55]
Ober-Olm (rye) 78.0 120.0 128.0 180.0 Hex/Ac NA [55]
Ober-Olm (rye) 35.0 30.0 48.0 70.0 Hex/Ac NA [55]
Ober-Olm (rye) 80.0 60.0 110.0 145.0 Hex/Ac NA [55]
Agricultural soil (Iran) 284.0 NT 2.8 NT MetOH NA [309]
Aligarh Soil 1 62.0 67.0 ND ND MetOH NA [60]
Aligarh Soil 1 64.0 79.0 ND ND ACN NA [60]
Aligarh Soil 1 52.0 49.0 ND ND Acetone NA [60]
II. Urban and/or residential sites
River edge (urban/industrial) ND 540.0 ND 2450.0 DCM NA [432]
Marsh (urban loaction) ND 260.0 ND 1800.0 DCM NA [432]
River edge (urban/industrial) ND 170.0 ND 620.0 DCM NA [432]
River edge (urban/residential) ND 130.0 ND 1000.0 DCM NA [432]
River edge (urban location) ND 80.0 ND 580.0 DCM NA [432]
River edge (urban/industrial) ND 70.0 ND 230.0 DCM NA [432]
River edge (urban/industrial) ND 60.0 ND 240.0 DCM NA [432]
River edge (urban/industrial) ND 60.0 ND 160.0 DCM NA [432]
Grassland (residential) NT 61.0 NT 88.0 DCM NA PAHs PAHs: 0.7 [39]
Woodland (urban/industrial) NT 51.0 NT 106.0 DCM NA PAHs PAHs: 54.5 [39]
Grassland (residential) NT 50.0 NT 93.0 DCM NA PAHs PAHs: 7.0 [39]
Park (urban) NT 48.0 NT 79.0 DCM NA PAHs PAHs: 3.8 [39]
Grassland (urban/industrial) NT 43.0 NT 82.0 DCM NA PAHs PAHs: 1.0 [39]
Pasture (urban/industrial) NT 41.0 NT 119.0 DCM NA PAHs PAHs: 5.4 [39]
Roadside (urban/residential) NT 20.0 NT 87.0 DCM NA PAHs PAHs: 8.3 [39]
Industrial site (background) NA 229.0 NA NA DCM/MetOH NA Creosote [45]
Urban park (Nara) ND 30.0 ND ND MetOH 0.57 Nitroarenes [124]
Urban park (Mino) ND 81.0 ND ND MetOH 1.07 Nitroarenes [124]
Urban park (Ibaraki) 84.0 190.0 88.0 140.0 MetOH 0.85 Nitroarenes [124]
Urban park (Sumiyoshi-ku) 5600.0 5900.0 600.0 2400.0 MetOH 0.75 Nitroarenesj 1,6-DNP:
1.9 � 10�3,
1,8-DNP:
2.2 � 10�3
[124]
Urban park (Minato-ku) 5900.0 5500.0 530.0 2500.0 MetOH 0.91 Nitroarenes 1,6-DNP:
1.7 � 10�3,
1,8-DNP:
2.2 � 10�3
[124]
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
0
Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
ppm dry weightc
Urban Park (Takarazuka) ND 58.0 ND ND MetOH 0.18 renes [124]
Urban soil (Azerbaijan 1) NT 84.0 NT NT Ac/Hex 0.84 PAHs: 7.3 [56]
Urban soil (Azerbaijan 2) NT 19.0 NT NT Ac/Hex 3.36 PAHs: 51.3 [56]
Urban soil (Azerbaijan 3) NT 21.0 NT NT Ac/Hex 2.22 PAHs: 11.7 [56]
Urban soil (Azerbaijan 5) NT 57.0 NT NT Ac/Hex 0.61 [56]
Urban soil (Azerbaijan 6) NT 23.0 NT NT Ac/Hex 1.66 PAHs: 8.79 [56]
Urban soil (Azerbaijan 7) NT 158.0 NT NT Ac/Hex 0.60 PAHs: 2.93 [56]
Urban soil (Azerbaijan 9) NT 8.0 NT NT Ac/Hex 8.38 PAHs: 11.72 [56]
Urban soil (Azerbaijan 10) NT 15.0 NT NT Ac/Hex 5.18 PAHs: 2.93 [56]
Urban soil (Sendai City) 77.0 408.0 241.0 347.0 NA NA carbons [431]
Urban soil (Kitakyushu city) 440.0 800.0 NT NT NA NA Hs PAHs: 0.96 [119]
Urban soil (Kyushu hway) 530.0 820.0 NT NT NA NA Hs PAHs: 0.34 [119]
Urban soil (Chugoku tunnel) 2600.0 6100.0 NT NT NA NA Hs PAHs: 0.28 [119]
Urban soil (Gyutozan) 2420.0 5680.0 NT NT NA NA Hs PAHs: 0.52 [119]
Urban soil (Kakei-nishi) 3860.0 8980.0 NT NT NA NA Hs PAHs: 0.35 [119]
Urban soil (Kakei-higashi) 2800.0 7030.0 NT NT NA NA Hs PAHs: 0.13 [119]
Urban soil (Yoneyama) 817.0 1590.0 NT NT NA NA Hs PAHs: 0.11 [119]
Mainz-Finthem Motorway 65.0 0.503 395.0 361.0 DCM/Ac/Tol:DEK 4.50 [116]
Mainz-Finthem Motorway 89.0 0.510 533.0 756.0 DCM/Ac/Tol:DEK 7.78 [116]
Mainz-Finthem Motorway 472.0 1.890 4890.0 3427.0 DCM/Ac/Tol:DEK 4.29 PAHs: 17.3 [116]
A60, exit Groberg 450.0 448.0 NT NT DCM/Ac/Tol:DEK NA [116]
A60, exit Grobberg 1170.0 292.0 NT NT DCM/Ac/Tol:DEK NA [116]
Saarstrabe near Mainz ND 564.0 NT NT DCM/Ac/Tol:DEK NA [116]
Saarstrabe near Mainz 1060.0 600.0 NT NT DCM/Ac/Tol:DEK NA [116]
Bundesstrabe 9 near Labenheim ND 1240.0 NT NT DCM/Ac/Tol:DEK NA [116]
Bundesstrabe 9 near Labenheim 600.0 450.0 NT NT DCM/Ac/Tol:DEK NA [116]
Bundesstrabe 9 near Labenheim 181.0 474.0 NT NT DCM/Ac/Tol:DEK NA [116]
Landstrabe L431 1030.0 1700.0 NT NT DCM/Ac/Tol:DEK NA [116]
Landstrabe L431 1100.0 519.0 NT NT DCM/Ac/Tol:DEK NA [116]
Landstrabe L431 1100.0 1850.0 NT NT DCM/Ac/Tol:DEK NA [116]
Landstrabe L422 800.0 1013.0 NT NT DCM/Ac/Tol:DEK NA [116]
Landstrabe L422 138.0 45.0 NT NT DCM/Ac/Tol:DEK NA [116]
Name
Nitroa
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
Hydro
Tar/PA
Tar/PA
Tar/PA
Tar/PA
Tar/PA
Tar/PA
Tar/PA
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
PAHs
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
1
Landstrabe L422 32.0 190.0 NT NT DCM/Ac/Tol:DEK NA PAHs [116]
Tokyo soil (commercial) 226.0 325.0 NT NT Benzene/EtOH NA PAHs BaPk: 0.20 [327]
Tokyo soil (residential) 147.0 290.0 NT NT Benzene/EtOH NA PAHs BaP: 0.16 [327]
Tokyo Shinagawa-ku 319.0 NT NT NT MetOH NA PAHs, DNPs DNPs:
0.18 � 10�3[52]
Higashimurayama 438.0 NT NT NT MetOH NA s, DNPs DNPs:
0.05 � 10�3[52]
Hachioji 380.0 NT NT NT MetOH NA s, DNPs DNPs:
0.07 � 10�3[52]
Nagoya 180.0 NT NT NT MetOH NA s, DNPs DNPs:
0.04 � 10�3[52]
Gifu 260.0 NT NT NT MetOH NA s, DNPs DNPs:
0.21 � 10�3[52]
Uji 3300.0 NT NT NT MetOH NA s, DNPs DNPs:
1.81 � 10�3[52]
Osaka Sumiyoshi-ku 9780.0 NT NT NT MetOH NA s, DNPs DNPs:
9.40 � 10�3[52]
Higashiosaka 248.0 NT NT NT MetOH NA s, DNPs DNPs:
0.12 � 10�3[52]
Sapporo 1 360.0 582.0 383.0 2092.0 MetOH NA s, DNPs [53]
Sapporo 2 606.0 825.0 594.0 1793.0 MetOH NA s, DNPs [53]
Sapporo 3 381.0 502.0 414.0 1641.0 MetOH NA s, DNPs [53]
Sapporo 4 214.0 306.0 225.0 732.0 MetOH NA s, DNPs [53]
Sapporo 5 512.0 664.0 516.0 752.0 MetOH NA s, DNPs [53]
Sapporo 6 214.0 582.0 320.0 357.0 MetOH NA s, DNPs [53]
Sapporo 7 912.0 1076.0 772.0 912.0 MetOH NA s, DNPs [53]
Sapporo 8 472.0 615.0 508.0 448.0 MetOH NA s, DNPs [53]
Muroran 1 194.0 428.0 424.0 700.0 MetOH NA s, DNPs [53]
Muroran 2 404.0 1368.0 876.0 2420.0 MetOH NA s, DNPs [53]
Muroran 3 366.0 2135.0 558.0 5259.0 MetOH NA s, DNPs [53]
Muroran 4 624.0 2400.0 616.0 2676.0 MetOH NA s, DNPs [53]
Muroran 5 372.0 1952.0 560.0 6360.0 MetOH NA s, DNPs [53]
Muroran 6 130.0 135.0 286.0 294.0 MetOH NA s, DNPs [53]
Asahikawa 1 206.0 336.0 265.0 741.0 MetOH NA s, DNPs [53]
Asahikawa 2 488.0 704.0 668.0 968.0 MetOH NA s, DNPs [53]
Asahikawa 3 454.0 570.0 790.0 2054.0 MetOH NA s, DNPs [53]
Asahikawa 4 129.0 132.0 177.0 726.0 MetOH NA s, DNPs [53]
Asahikawa 5 191.0 186.0 219.0 772.0 MetOH NA s, DNPs [53]
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
PAH
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
2
Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
Kushiro 1 170.0 194.0 261.0 648.0 MetOH NA PAHs, Ps [53]
Kushiro 2 212.0 210.0 240.0 336.0 MetOH NA PAHs, Ps [53]
Kushiro 3 177.0 285.0 265.0 350.0 MetOH NA PAHs, Ps [53]
Kushiro 4 114.0 254.0 239.0 414.0 MetOH NA PAHs, Ps [53]
Kushiro 5 159.0 154.0 120.0 370.0 MetOH NA PAHs, Ps [53]
Tokyo 1 1240.0 885.0 937.0 735.0 MetOH NA PAHs, Ps [53]
Tokyo 2 324.0 335.0 252.0 281.0 MetOH NA PAHs, Ps [53]
Tokyo 3 596.0 648.0 242.0 501.0 MetOH NA PAHs, Ps [53]
Tokyo 4 508.0 588.0 743.0 300.0 MetOH NA PAHs, Ps [53]
Tokyo 5 425.0 528.0 279.0 581.0 MetOH NA PAHs, Ps [53]
Tokyo 6 282.0 445.0 198.0 357.0 MetOH NA PAHs, Ps [53]
Tokyo 7 2310.0 1300.0 1800.0 1390.0 MetOH NA PAHs, Ps [53]
Tokyo 8 303.0 435.0 372.0 227.0 MetOH NA PAHs, Ps [53]
Higashikurume 137.0 318.0 316.0 383.0 MetOH NA PAHs, Ps [53]
Kawasaki 315.0 431.0 732.0 496.0 MetOH NA PAHs, Ps [53]
Yokohama 1 345.0 403.0 428.0 727.0 MetOH NA PAHs, Ps [53]
Yokohama 2 1060.0 1310.0 539.0 989.0 MetOH NA PAHs, Ps [53]
Yokohama 3 204.0 252.0 141.0 477.0 MetOH NA PAHs, Ps [53]
Ebina 243.0 347.0 693.0 251.0 MetOH NA PAHs, Ps [53]
Warabi 440.0 755.0 511.0 568.0 MetOH NA PAHs, Ps [53]
Omiya 934.0 995.0 587.0 1300.0 MetOH NA PAHs, Ps [53]
Kumagaya 395.0 383.0 389.0 598.0 MetOH NA PAHs, Ps [53]
Funabashi 184.0 247.0 268.0 484.0 MetOH NA PAHs, Ps [53]
Tsukuba 468.0 444.0 574.0 445.0 MetOH NA PAHs, Ps [53]
Maebashi 195.0 310.0 84.0 391.0 MetOH NA PAHs, Ps [53]
Nagoya 1 139.0 261.0 29.0 80.0 MetOH NA PAHs, Ps [53]
Nagoya 2 61.0 363.0 151.0 154.0 MetOH NA PAHs, Ps [53]
Nagoya 3 213.0 237.0 285.0 226.0 MetOH NA PAHs, Ps [53]
Nagoya 4 629.0 949.0 613.0 607.0 MetOH NA PAHs, Ps [53]
Nagoya 5 168.0 493.0 240.0 339.0 MetOH NA PAHs, Ps [53]
Nagoya 6 275.0 645.0 223.0 340.0 MetOH NA PAHs, Ps [53]
Ogaki 1 56.0 93.0 149.0 69.0 MetOH NA PAHs, Ps [53]
Ogaki 2 347.0 560.0 603.0 445.0 MetOH NA PAHs, Ps [53]
Ogaki 3 341.0 1060.0 861.0 875.0 MetOH NA PAHs, Ps [53]
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
DN
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
3
Okazaki 1 461.0 316.0 181.0 148.0 MetOH NA PAHs, DNPs [53]
Okazaki 2 180.0 648.0 587.0 635.0 MetOH NA PAHs, DNPs [53]
Okazaki 3 376.0 587.0 319.0 619.0 MetOH NA PAHs, DNPs [53]
Hekinan 1 168.0 136.0 139.0 147.0 MetOH NA PAHs, DNPs [53]
Hekinan 2 26300.0 10400.0 4600.0 6720.0 MetOH NA PAHs, NPs [53]
Ichinomiya 1 477.0 805.0 398.0 539.0 MetOH NA PAHs, NPs [53]
Ichinomiya 2 373.0 665.0 408.0 424.0 MetOH NA PAHs, NPs [53]
Nakatsugawa 1 141.0 159.0 91.0 63.0 MetOH NA PAHs, NPs [53]
Nakatsugawa 2 147.0 408.0 248.0 224.0 MetOH NA PAHs, NPs [53]
Shinshiro 1 256.0 253.0 117.0 144.0 MetOH NA PAHs, NPs [53]
Shinshiro 2 440.0 597.0 239.0 299.0 MetOH NA PAHs, NPs [53]
Toyohashi 1 357.0 325.0 64.0 123.0 MetOH NA PAHs, NPs [53]
Toyohashi 2 117.0 304.0 159.0 132.0 MetOH NA PAHs, NPs [53]
Inasa 220.0 312.0 147.0 467.0 MetOH NA PAHs, NPs [53]
Kariya 379.0 445.0 232.0 320.0 MetOH NA PAHs, NPs [53]
Kaizu 339.0 608.0 385.0 301.0 MetOH NA PAHs, NPs [53]
Moriyama 46.0 98.0 ND 47.0 MetOH NA PAHs, NPs [53]
Kusatsu 66.0 101.0 ND 0.0 MetOH NA PAHs, NPs [53]
Otsu 1 256.0 629.0 ND 120.0 MetOH NA PAHs, NPs [53]
Otsu 2 52.0 92.0 ND 69.0 MetOH NA PAHs, NPs [53]
Kyoto 1 83.0 ND 0.0 MetOH NA PAHs, NPs [53]
Kyoto 2 87.0 173.0 ND 55.0 MetOH NA PAHs, NPs [53]
Gose 98.0 260.0 ND 0.0 MetOH NA PAHs, NPs [53]
Nara ND 34.0 ND 0.0 MetOH NA PAHs, NPs [53]
Minoo 84.0 95.0 51.0 0.0 MetOH NA PAHs, NPs [53]
Ibaraki 64.0 138.0 92.0 132.0 MetOH NA PAHs, NPs [53]
Takatsuki 325.0 830.0 86.0 201.0 MetOH NA PAHs, NPs [53]
Osaka 1 6735.0 6515.0 585.0 2672.0 MetOH NA PAHs, NPs DNPs:
4.97 � 10�3[53]
Osaka 2 5963.0 4255.0 527.0 2397.0 MetOH NA PAHs, NPs DNPs:
5.94 � 10�3[53]
Osaka 3 225.0 702.0 ND 141.0 MetOH NA PAHs, NPs [53]
Tararazuka ND 32.0 ND 0.0 MetOH NA PAHs, NPs [53]
Ashiya 103.0 272.0 ND 153.0 MetOH NA PAHs, NPs [53]
Nishinomiya 1532.0 3965.0 144.0 745.0 MetOH NA PAHs, NPs [53]
Amagasaki 380.0 884.0 ND 284.0 MetOH NA PAHs, NPs [53]
Kobe 1 10167.0 4302.0 720.0 3275.0 MetOH NA PAHs, NPs DNPs:
5.54 � 10�3[53]
Kobe 2 162.0 552.0 82.0 67.0 MetOH NA PAHs, NPs [53]
Kitakyushu 1 370.0 419.0 155.0 470.0 MetOH NA PAHs, NPs [53]
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
4
Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
Kitakyushu 2 581.0 726.0 367.0 598.0 MetOH NA PAHs, NPs [53]
Dazaifu 1 331.0 600.0 127.0 509.0 MetOH NA PAHs, NPs [53]
Dazaifu 2 176.0 287.0 111.0 251.0 MetOH NA PAHs, NPs [53]
Fukuoka 1 381.0 571.0 308.0 457.0 MetOH NA PAHs, NPs [53]
Fukuoka 2 330.0 388.0 153.0 486.0 MetOH NA PAHs, NPs [53]
Fukuoka 3 224.0 458.0 111.0 480.0 MetOH NA PAHs, NPs [53]
Fukuoka 4 348.0 449.0 205.0 436.0 MetOH NA PAHs, NPs [53]
Kurume 1 349.0 598.0 169.0 519.0 MetOH NA PAHs, NPs [53]
Kurume 2 284.0 308.0 118.0 327.0 MetOH NA PAHs, NPs [53]
Kurume 3 84.0 252.0 36.0 159.0 MetOH NA PAHs, NPs [53]
Yame 1 409.0 645.0 346.0 476.0 MetOH NA PAHs, NPs [53]
Yame 2 272.0 595.0 276.0 485.0 MetOH NA PAHs, NPs [53]
Tosu 1 421.0 982.0 198.0 537.0 MetOH NA PAHs, NPs [53]
Tosu 2 393.0 621.0 308.0 663.0 MetOH NA PAHs, NPs [53]
Ohmuta 1 417.0 661.0 302.0 517.0 MetOH NA PAHs, NPs [53]
Ohmuta 2 782.0 1405.0 586.0 806.0 MetOH NA PAHs, NPs [53]
Ohmuta 3 648.0 1491.0 481.0 686.0 MetOH NA PAHs, NPs [53]
Ohmuta 4 549.0 922.0 435.0 657.0 MetOH NA PAHs, NPs [53]
Ohmuta 5 666.0 669.0 639.0 554.0 MetOH NA PAHs, NPs [53]
Ohmuta 6 412.0 792.0 330.0 662.0 MetOH NA PAHs, NPs [53]
Hekinan 1 NT 34300.0 NT NT MetOH NA PAHs, NPs DNPs:
0.12 � 10�3[51]
Hekinan 2 NT 46800.0 NT NT MetOH NA PAHs, NPs DNPs:
15.67 � 10�3[51]
Kurume site 1 528.0 679.0 ND 424.0 MetOH NA PAHs, etals PAHs: 0.60 [122]
Kurume site 2 327.0 394.0 ND 302.0 MetOH NA PAHs, etals PAHs: 0.21 [122]
Kurume site 3 95.0 79.0 ND ND MetOH NA PAHs, etals PAHs: 0.03 [122]
Kurume site 4 151.0 71.0 ND ND MetOH NA PAHs, etals PAHs: 0.008 [122]
Kurume site 5 221.0 125.0 ND ND MetOH NA PAHs, etals PAHs: 0.25 [122]
Kurume site 6 541.0 649.0 198.0 343.0 MetOH NA PAHs, etals PAHs: 1.07 [122]
Kurume site 7 903.0 808.0 195.0 386.0 MetOH NA PAHs, etals PAHs: 0.69 [122]
Kurume site 8 598.0 750.0 231.0 437.0 MetOH NA PAHs, etals PAHs: 0.18 [122]
Kurume site 9 289.0 349.0 ND 203.0 MetOH NA PAHs, etals PAHs: 0.14 [122]
Kurume site 10 463.0 604.0 367.0 550.0 MetOH NA PAHs, etals PAHs: 0.10 [122]
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
M
M
M
M
M
M
M
M
M
M
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
5
Kurume site 11 721.0 840.0 234.0 366.0 MetOH NA PAHs, Metals PAHs: 1.11 [122]
Kurume site 12 432.0 570.0 ND 306.0 MetOH NA PAHs, Metals PAHs: 0.15 [122]
Kurume site 13 724.0 279.0 ND ND MetOH NA PAHs, Metals PAHs: 0.04 [122]
Kokagun Kokacho 81.0 81 ND 120.0 MetOH NA PAHs, DNPs [50]
Kokagun Minakuchicho 193.0 470.0 81.0 193.0 MetOH NA PAHs, NPs [50]
Kusatu 103.0 256.0 ND 161.0 MetOH NA PAHs, NPs [50]
Moriyama 1 194.0 483.0 ND 212.0 MetOH NA PAHs, NPs [50]
Moriyama 2 120.0 158.0 ND 159.0 MetOH NA PAHs, NPs [50]
Omihachiman 47.0 81.0 ND 81.0 MetOH NA PAHs, NPs [50]
Otsu 158.0 354.0 77.0 227.0 MetOH NA PAHs, NPs [50]
Kameoka 30.0 118.0 122.0 162.0 MetOH NA PAHs, NPs [50]
Kyoto Fushimi-ku 288.0 142.0 109.0 156.0 MetOH NA PAHs, NPs [50]
Kyoto Kamigyo-ku 294.0 931.0 230.0 1067.0 MetOH NA PAHs, NPs [50]
Kyoto Ukyo-ku 258.0 368.0 184.0 244.0 MetOH NA PAHs, NPs [50]
Kyoto Yamashina-ku 2325.0 2727.0 501.0 1139.0 MetOH NA PAHs, NPs [50]
Nagaokakyo 3000.0 877.0 314.0 384.0 MetOH NA PAHs, NPs [50]
Sonobecho 280.0 525.0 117.0 277.0 MetOH NA PAHs, NPs [50]
Uji 4797.0 2829.0 462.0 1674.0 MetOH NA PAHs, NPs DNPs:
2.84 � 10�3[50]
Nara 71.0 91.0 70.0 133.0 MetOH NA PAHs, NPs [50]
Tenri 154.0 615.0 71.0 352.0 MetOH NA PAHs, NPs [50]
Yamatokoriyama 252.0 483.0 119.0 373.0 MetOH NA PAHs, NPs [50]
Ibaraki 1 202.0 541.0 ND 143.0 MetOH NA PAHs, NPs [50]
Ibaraki 2 1104.0 4256.0 274.0 960.0 MetOH NA PAHs, NPs [50]
Izumi 1 1598.0 2032.0 343.0 1236.0 MetOH NA PAHs, NPs DNPs:
0.20 � 10�3[50]
Izumi 2 146.0 309.0 ND 111.0 MetOH NA PAHs, NPs [50]
Kadoma 448.0 309.0 ND 259.0 MetOH NA PAHs, NPs [50]
Kishiwada 1627.0 10898.0 607.0 2057.0 MetOH NA PAHs, NPs DNPs:
0.03 � 10�3[50]
Moriguchi 1979.0 5619.0 239.0 1325.0 MetOH NA PAHs, NPs [50]
Neyagawa 534.0 776.0 ND 432.0 MetOH NA PAHs, NPs [50]
Osaka Abeno-ku 3056.0 3152.0 411.0 1271.0 MetOH NA PAHs, NPs DNPs:
3.07 � 10�3[50]
Osaka Chuo-ku 146.0 260.0 ND 212.0 MetOH NA PAHs, NPs [50]
Osaka Fukushima-ku 579.0 749.0 92.0 664.0 MetOH NA PAHs, NPs DNPs:
0.37 � 10�3[50]
Osaka Higashinari-ku 1627.0 1488.0 320.0 976.0 MetOH NA PAHs, NPs DNPs:
0.11 � 10�3[50]
Osaka Higashisumiyoshi-ku 130.0 160.0 235.0 190.0 MetOH NA PAHs, NPs DNPs:
0.05 � 10�3[50]
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
6
Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
Osaka Higashiyodogawa-ku 2710.0 1296.0 116.0 969.0 MetOH NA PAHs, DNPs DNPs:
2.24 � 10�3[50]
Osaka Ikuno-ku 334.0 716.0 107.0 1271.0 MetOH NA PAHs, DNPs DNPs:
0.28 � 10�3[50]
Osaka Joto-ku 1136.0 2314.0 400.0 760.0 MetOH NA PAHs, DNPs DNPs:
0.66 � 10�3[50]
Osaka Kita-ku 4232.0 1842.0 109.0 2156.0 MetOH NA PAHs, DNPs DNPs:
2.59 � 10�3[50]
Osaka Konohana-ku 229.0 747.0 186.0 528.0 MetOH NA PAHs, DNPs [50]
Osaka Minato-ku 5963.0 2981.0 307.0 1515.0 MetOH NA PAHs, DNPs DNPs:
5.39 � 10�3[50]
Osaka Miyakojima-ku 110.0 472.0 ND 603.0 MetOH NA PAHs, DNPs [50]
Osaka Naniwa-ku 224.0 423.0 115.0 491.0 MetOH NA PAHs, DNPs DNPs:
0.10 � 10�3[50]
Osaka Nishi-ku 1420.0 3430.0 473.0 1900.0 MetOH NA PAHs, DNPs DNPs:
0.32 � 10�3[50]
Osaka Nishinari-ku 224.0 947.0 ND 528.0 MetOH NA PAHs, DNPs [50]
Osaka Suminoe-ku 6075.0 7085.0 980.0 1869.0 MetOH NA PAHs, DNPs DNPs:
2.97 � 10�3[50]
Osaka Sumiyoshi-ku 1595.0 2629.0 765.0 1349.0 MetOH NA PAHs, DNPs [50]
Osaka Tennoji-ku 129.0 458.0 ND 293.0 MetOH NA PAHs, DNPs [50]
Osaka Tsurumi-ku 112.0 544.0 ND 384.0 MetOH NA PAHs, DNPs [50]
Sakai 1 3073.0 3659.0 448.0 1957.0 MetOH NA PAHs, DNPs DNPs:
2.41 � 10�3[50]
Sakai 2 1302.0 4804.0 158.0 1027.0 MetOH NA PAHs, DNPs DNPs:
0.04 � 10�3[50]
Sakai 3 4092.0 4656.0 616.0 2941.0 MetOH NA PAHs, DNPs DNPs:
0.90 � 10�3[50]
Sakai 4 1208.0 2352.0 212.0 458.0 MetOH NA PAHs, DNPs [50]
Suita 425.0 630.0 ND 557.0 MetOH NA PAHs, DNPs DNPs:
0.04 � 10�3[50]
Takatsuki 2408.0 1552.0 338.0 830.0 MetOH NA PAHs, DNPs DNPs:
0.69 � 10�3[50]
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
7
Takaishi 2581.0 2128.0 407.0 1779.0 MetOH NA PAHs, DNPs DNPs:
0.91 � 10�3[50]
Toyonaka 2768.0 2755.0 379.0 1000.0 MetOH NA PAHs, DNPs [50]
Amagasaki 691.0 1197.0 106.0 507.0 MetOH NA PAHs, DNPs DNPs:
.33 � 10�3[50]
Itami 382.0 529.0 86.0 366.0 MetOH NA PAHs, DN [50]
Kobe Nada-ku 755.0 642.0 71.0 363.0 MetOH NA PAHs, DN [50]
Kobe Nagata-ku 496.0 1168.0 181.0 1392.0 MetOH NA PAHs, DN NPs:
.32 � 10�3[50]
Nishinomiya 183.0 775.0 128.0 459.0 MetOH NA PAHs, DN [50]
Kainan 1 117.0 443.0 ND 189.0 MetOH NA PAHs, DN [50]
Kainan 2 403.0 2239.0 186.0 741.0 MetOH NA PAHs, DN [50]
Wakayama 1 744.0 2057.0 ND 542.0 MetOH NA PAHs, DN [50]
Wakayama 2 545.0 1182.0 104.0 415.0 MetOH NA PAHs, DN [50]
III. Highly contaminated industrial sites (including superfund sites)
Superfund soil (3)l NA 376237.6 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (10) NA 242038.2 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (5) NA 238993.7 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (2) NA 136200.7 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (4) NA 121019.1 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (9) NA 116564.4 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (10) NA 104683.2 NA NA Cyclohex NA PAHs AHs: 35–530 [46]
Superfund soil (7) NA 88993.0 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (6) NA 77079.1 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (5) NA 37076.7 NA NA Cyclohex NA PAHs AHs: 35–530 [46]
Superfund soil (9) NA 33989.3 NA NA Cyclohex NA PAHs AHs: 35–530 [46]
Superfund soil (3) NA 31746.0 NA NA Cyclohex NA PAHs AHs: 35–530 [46]
Superfund soil (8) NA 27818.4 NA NA DCM NA PAHs AHs: 35–530 [46]
Superfund soil (6) NA 27676.6 NA NA Cyclohex NA PAHs AHs: 35–530 [46]
Superfund soil (8) NA 16740.1 NA NA Cyclohex NA PAHs AHs: 35–530 [46]
Superfund soil (2) NA 13446.6 NA NA Cyclohex NA PAHs AHs: 35–530 [46]
Wood-preserving site NA 23518.0 NA NA DCM/MetOH 53.90 PAHs, PC
PCDDsmCDFs + PCDDs: �2 � 10�3
AHs: 12267.0
[45]
Amended soil (creosote) 367.0 23480.0 NA 21475.0 DCM 134.20 Creosote, [42]
0
Ps
Ps
Ps D
0
Ps
Ps
Ps
Ps
Ps
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
DFs, P
P
PAHs
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
8
Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
Amended soil (creosote) 140.0 9921.0 NA 5632.0 DCM 135.00 Creosote, PAHs [42]
Amended soil (petroleum) NA 4804.0 NA NA DCM 11.38 PAHs [166]
Amended soil (petroleum) NA 3856.0 NA NA DCM 14.18 PAHs [166]
Amended soil (petroleum) NA 2155.0 NA NA DCM 9.58 Hydrocarbons PAHs [166]
Amended soil (petroleum) NA 1459.0 NA NA DCM 4.33 Hydrocarbons PAHs [166]
Industrial site (DCT 24) 734.0 664.0 NA NA DCM 1.26 Dyes, Pesticides 2,3,6-TCBAn:
16.0 ABN-X:
16.3
[47]
Industrial site (DCT 38) 438.0 456.0 NA NA DCM 1.78 Dyes, Pesticides 2-M-5-NBA:
0.1 ABN-X:
1.42-NA: 0.3
[47]
Industrial site (DCT 24) 352.0 412.0 NA NA MetOH 0.28 Dyes, Pesticides 2,3,6-TCBA:
16.0 ABN-X:
16.3
[47]
Industrial site (DCT 31) 197.0 263.0 NA NA DCM 0.28 Dyes, Pesticides [47]
Industrial site (DCT 25) 143.0 161.0 NA NA MetOH 0.075 Dyes, Pesticides [47]
Industrial site (DCT 26) 170.0 135.0 NA NA DCM 0.074 Dyes, Pesticides [47]
Industrial site (DCT 26) 124.0 132.0 NA NA MetOH 0.050 Dyes, Pesticides [47]
Industrial site (DCT 32) 81.0 94.0 NA NA MetOH 0.029 Dyes, Pesticides [47]
Industrial site (DCT 38) 101.0 86.0 NA NA MetOH 0.037 Dyes, Pesticides 2-M-5-NBA:
0.1 ABN-X:
1.42-NA: 0.3
[47]
Industrial site (DCT 25) 40.0 42.0 NA NA DCM 0.014 Dyes, Pesticides [47]
Industrial site (DCT 32) 15.0 20.0 NA NA DCM 0.008 Dyes, Pesticides [47]
Industrial site (DCT 31) 2.0 3.0 NA NA MetOH 0.001 Dyes, Pesticides [47]
Industrial site (Soil 505)o ND 194.0 NT NT MetOH 2.40 Solvent, metals, paint sludge [114]
Industrial site (Soil 506) 80.0 ND NT NT MetOH 1.27 Solvent, metals, paint sludge [114]
Industrial site (Soil 302) 2449.0 ND NT NT MetOH 5.43 Solvent, metals, paint sludge [114]
Industrial site (Soil 302) 2370.0 ND NT NT DCM 5.43 Solvent, metals, paint sludge [114]
Industrial site (Soil 303) 1204.0 ND NT NT MetOH 3.22 Solvent, metals, paint sludge [114]
Industrial site (Soil 303) 2394.0 ND NT NT DCM 4.45 Solvent, metals, paint sludge [114]
Industrial site (Soil 204) 1380.0 ND NT NT DCM 11.90 Solvent, metals, paint sludge [114]
Industrial site (Soil 402) 1356.0 ND NT NT DCM 7.14 Solvent, metals, paint sludge [114]
Industrial site (Soil 502) 741.0 ND NT NT MetOH 4.49 Solvent, metals, paint sludge [114]
Industrial site (Soil 502) 532.0 ND NT NT DCM 9.50 Solvent, metals, paint sludge [114]
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
30
9
Industrial site (Soil 005) 523.0 105.0 NT NT MetOH 5.53 Solvent, metals, paint sludge [114]
Industrial site (Soil 005) 1192.0 162.0 NT NT DCM 10.12 Solvent, metals, paint sludge [114]
Industrial site (Soil 503) ND 77.0 NT NT DCM 1.75 Solvent, metals, paint sludge [114]
Industrial site (Soil 110) 791.0 22.0 NT NT MetOH 11.30 Solvent, metals, paint sludge [114]
Industrial site (Soil 110) 1041.0 ND NT NT DCM 25.39 Solvent, metals, paint sludge [114]
PCB contaminated soil 122.0 197.0 ND ND DCM 3.58 PCBs Aroclor
1260: � 2000
[41]
PCB contaminated soil 58.0 120.0 ND ND DCM 1.36 PCBs Aroclor
1260: � 2000
[41]
Amended soil (creosote) 25.0 782.5 NT NT DCM/MetOH NA Creosote [64]
Amended soil (refinery) ND 329.0 NT NT DCM/MetOH NA PAHs [64]
Industrial site (Soil 201) ND 16.0 NT NT DCM 0.20 PAHs [118]
Industrial site (Soil 001) ND 28183.0 NT NT DCM 276.30 PAHs [118]
Industrial site (Soil 002) ND 175195.0 NT NT DCM 141.29 PAHs [118]
Industrial site (Soil 003) ND 178488.0 NT NT DCM 815.01 PAHs [118]
Industrial site (Soil 102) ND 38622.0 NT NT DCM 495.15 PAHs [118]
Industrial site (Soil 103) ND 28498.0 NT NT DCM 459.64 PAHs [118]
Industrial site (Soil 104) ND 72518.0 NT NT DCM 659.25 PAHs [118]
Industrial site (Soil 105) ND 4890.0 NT NT DCM 116.43 PAHs [118]
Industrial site (Soil 108) ND 51403.0 NT NT DCM 634.61 PAHs [118]
Industrial site (Soil 202) ND 181847.0 NT NT DCM 790.64 PAHs [118]
Industrial site (Soil 203) ND 161892.0 NT NT DCM 700.83 PAHs PAHs: 2168.0 [118]
Industrial site (Soil 204) ND 206039.0 NT NT DCM 865.71 PAHs PAHs: 2533.0 [118]
Industrial site (Soil 205) ND 80119.0 NT NT DCM 340.93 PAHs PAHs: 1802.0 [118]
Industrial site (Soil 206) ND 252877.0 NT NT DCM 909.63 PAHs [118]
Industrial site (Soil 201) 45.0 32.0 NT NT MetOH 0.46 PAHs [118]
Industrial site (Soil 001) ND 24919.0 NT NT MetOH 307.64 PAHs [118]
Industrial site (Soil 002) ND 5502.0 NT NT MetOH 250.09 PAHs [118]
Industrial site (Soil 003) ND 14786.0 NT NT MetOH 122.20 PAHs [118]
Industrial site (Soil 102) ND 5749.0 NT NT MetOH 6.78 PAHs [118]
Industrial site (Soil 103) ND 758.0 NT NT MetOH 12.23 PAHs [118]
Industrial site (Soil 104) ND 1801.0 NT NT MetOH 9.38 PAHs [118]
Industrial site (Soil 105) ND 337.0 NT NT MetOH 2.81 PAHs [118]
Industrial site (Soil 108) ND 24637.0 NT NT MetOH 13.47 PAHs [118]
Industrial site (Soil 202) ND 4322.0 NT NT MetOH 6.32 PAHs [118]
Industrial site (Soil 203) ND 9536.0 NT NT MetOH 12.94 PAHs PAHs: 581.0 [118]
Industrial site (Soil 204) ND 4654.0 NT NT MetOH 5.16 PAHs PAHs: 443.0 [118]
Industrial site (Soil 205) ND 7304.0 NT NT MetOH 9.40 PAHs PAHs: 895.0 [118]
Industrial site (Soil 206) ND 10222.0 NT NT MetOH 17.84 PAHs [118]
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0Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
Amended soil (sewage sludge) 246.0 105.0 NT NT DCM NA [179]
Amended soil (sewage sludge) 106.3 70.0 NT NT DCM NA [179]
Amended soil (sewage sludge) 234.0 98.0 NT NT DCM NA [179]
Amended soil (sewage sludge) 215.0 78.0 NT NT DCM NA [179]
Amended soil (sewage sludge) 130.0 75.0 NT NT MetOH NA [179]
Amended soil (sewage sludge) 23.0 15.0 NT NT MetOH NA [179]
Amended soil (sewage sludge) 80.0 58.0 NT NT MetOH NA [179]
Amended soil (sewage sludge) 58.0 43.0 NT NT MetOH NA [179]
Hazardous waste site (S2) ND 1.0 NT NT DCM 0.088 Solvents, t, PAHs PAHs: 1.0 [117]
Hazardous waste site (S4) ND 595.0 NT NT DCM 23.8 Solvents, t, PAHs [117]
Hazardous waste site (S6) ND 27.0 NT NT DCM NA Solvents, t, PAHs [117]
Hazardous waste site (S8) ND 37.0 NT NT DCM 0.55 Solvents, t, PAHs PAHs: 0.5 [117]
Hazardous waste site (S10) ND 71.0 NT NT DCM 0.46 Solvents, t, PAHs PAHs: 15.3 [117]
Hazardous waste site (S12) ND 2162.0 NT NT DCM 29.22 Solvents, t, PAHs PAHs: 74.0 [117]
Hazardous waste site (S14) ND 2.0 NT NT DCM 0.14 Backgrou [117]
Hazardous waste site (S4) ND 162.0 NT NT MetOH 2.19 Solvents, t, PAHs [117]
Hazardous waste site (S6) ND 44.0 NT NT MetOH 0.55 Solvents, t, PAHs [117]
Hazardous waste site (S8) ND 15.0 NT NT MetOH 0.71 Solvents, t, PAHs PAHs: 0.5 [117]
Hazardous waste site (S10) ND 32.0 NT NT MetOH 0.35 Solvents, t, PAHs PAHs: 15.3 [117]
Hazardous waste site (S12) ND 248.0 NT NT MetOH 1.77 Solvents, t, PAHs PAHs: 74.0 [117]
Hazardous waste site (S14) ND 8.0 NT NT MetOH 0.35 Backgrou [117]
Amended soil (sewage sludge) 154.0 81.0 NT NT DCM NA [156]
Amended soil (sewage sludge) 122.0 87.0 NT NT DCM NA [156]
Amended soil (sewage sludge) 105.0 147.0 NT NT MetOH NA [156]
Amended soil (sewage sludge) 170.0 426.0 NT NT MetOH NA [156]
Amended soil (sewage sludge) 310.0 371.0 NT NT DCM/MetOH NA [156]
Amended soil (sewage sludge) 422.0 509.0 NT NT DCM/MetOH NA [156]
Amended soil (creosote) 21.0 779.0 NT NT DCM/MetOH NA Creosote, [370]
Land treatment soil (005) ND 113.0 NT NT DCM 2.3 PAHs, PC PAHs: �126
PCP: �31
[115]
Land treatment soil (006) ND 59.0 NT NT DCM 2.8 PAHs, PC PAHs: �126
PCP: �31
[115]
Land treatment soil (007) ND 146.0 NT NT DCM 1.8 PAHs, PC PAHs: �126
PCP: �31
[115]
pain
pain
pain
pain
pain
pain
nd
pain
pain
pain
pain
pain
nd
PCP
P
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Land treatment soil (008) ND 174.0 NT NT DCM 2.1 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (009) ND 124.0 NT NT DCM 2.7 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (010) ND 67.0 NT NT DCM 1.2 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (011) ND 68.0 NT NT DCM 0.9 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (012) ND 67.0 NT NT DCM 1.2 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (013) ND 77.0 NT NT DCM 0.9 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (014) ND 97.0 NT NT DCM 2.1 PAHs, PCP PAHs: �279
PCP: �176
[115]
Land treatment soil (015) ND 114.0 NT NT DCM 3.0 PAHs, PCP PAHs: �279
PCP: �176
[115]
Land treatment soil (016) ND 71.0 NT NT DCM 0.6 PAHs, PCP PAHs: �279
PCP: �176
[115]
Land treatment soil (017) ND 66.0 NT NT DCM 0.4 Background [115]
Land treatment soil (023) ND 10.0 NT NT DCM 0.7 Background [115]
Land treatment soil (005) ND 21.0 NT NT MetOH 0.5 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (006) ND 331.0 NT NT MetOH 6.9 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (007) ND 77.0 NT NT MetOH 0.9 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (008) ND 43.0 NT NT MetOH 0.5 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (009) ND 34.0 NT NT MetOH 0.6 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (010) ND 5.0 NT NT MetOH 0.3 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (011) ND 46.0 NT NT MetOH 0.7 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (012) ND 47.0 NT NT MetOH 1.1 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (013) ND 70.0 NT NT MetOH 1.3 PAHs, PCP PAHs: �126
PCP: �31
[115]
Land treatment soil (014) ND 77.0 NT NT MetOH 1.5 PAHs, PCP PAHs: �279
PCP: �176
[115]
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Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
Land treatment soil (015) ND 102.0 NT NT MetOH 3.0 PAHs, PCP PAHs: �279
PCP: �176
[115]
Land treatment soil (016) ND 1.0 NT NT MetOH 0.5 PAHs, PCP PAHs: �279
PCP: �176
[115]
Land treatment soil (023) ND 8.0 NT NT MetOH 1.0 Background [115]
Land treatment soil (Cell 1) NA 328.0 NT NT DCM NA PAHs, PCP [56]
Land treatment soil (Cell 2) NA 594.0 NT NT DCM NA PAHs, PCP [56]
Land treatment soil (Cell 3) NA 440.0 NT NT DCM NA PAHs, PCP [56]
Land treatment soil (Cell 4) NA 380.0 NT NT DCM NA PAHs, PCP [56]
Land treatment soil (Cell 5) NA 475.0 NT NT DCM NA PAHs, PCP [56]
Land treatment soil (Cell 6) NA 523.0 NT NT DCM NA PAHs, PCP [56]
Land treatment soil (Cell 7) NA 978.0 NT NT DCM NA PAHs, PCP PAHs: 88.0 [56]
Land treatment soil (Cell 10) NA 337.0 NT NT DCM NA PAHs, PCP [56]
Land treatment soil (Cell 11) NA 333.0 NT NT DCM NA PAHs, PCP [56]
Land treatment soil (Cell 12) NA 311.0 NT NT DCM NA PAHs, PCP [56]
Munitions site (Soil 001) 32670 3277 NT NT DCM 9.93 Nitroaromatics [54]
Munitions site (Soil 002) 2935 371 NT NT DCM 1.16 Nitroaromatics [54]
Munitions site (Soil 003) 177559 ND NT NT DCM 105.7 Nitroaromatics [54]
Munitions site (Soil 004) 126008 19067 NT NT DCM 82.9 Nitroaromatics [54]
Munitions site (Soil 005) 58073 7743 NT NT DCM 25.8 Nitroaromatics [54]
Munitions site (Soil 006) ND 16.0 NT NT DCM 0.13 Background [54]
Munitions site (Soil 101) 119130 16281 NT NT DCM 39.7 Nitroaromatics [54]
Munitions site (Soil 102) 235165 22711 NT NT DCM 73.3 Nitroaromatics [54]
Munitions site (Soil 103) 20217 8959 NT NT DCM 1.2 Nitroaromatics [54]
Munitions site (Soil 104) 9531 1517 NT NT DCM 0.74 Nitroaromatics [54]
Munitions site (Soil 105) 288350 103806 NT NT DCM 115.3 Nitroaromatics [54]
Munitions site (Soil 106) 179516 22365 NT NT DCM 29.8 Nitroaromatics [54]
Munitions site (Soil 107) 279896 24194 NT NT DCM 47.4 Nitroaromatics [54]
Munitions site (Soil 108) 15532 2410 NT NT DCM 2.06 Nitroaromatics [54]
Munitions site (Soil 109) 1200 ND NT NT DCM 0.06 Background [54]
Munitions site (Soil 001) 19397 5726 NT NT MetOH 3.31 Nitroaromatics [54]
Munitions site (Soil 002) 7823 1522 NT NT MetOH 1.18 Nitroaromatics [54]
Munitions site (Soil 003) 50115 8327 NT NT MetOH 15.4 Nitroaromatics [54]
Munitions site (Soil 004) 36951 5297 NT NT MetOH 12.9 Nitroaromatics [54]
Munitions site (Soil 005) 105117 40900 NT NT MetOH 8.56 Nitroaromatics [54]
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Munitions site (Soil 101) 24897 1081 NT NT MetOH 3.86 Nitroaromatics [54]
Munitions site (Soil 102) 46982 11509 NT NT MetOH 2.78 Nitroaromatics [54]
Munitions site (Soil 103) 14523 1620 NT NT MetOH 1.78 Nitroaromatics [54]
Munitions site (Soil 104) 24286 4687 NT NT MetOH 0.72 Nitroaromatics [54]
Munitions site (Soil 105) 43737 14722 NT NT MetOH 2.39 Nitroaromatics [54]
Munitions site (Soil 106) 18681 7285 NT NT MetOH 0.73 Nitroaromatics [54]
Munitions site (Soil 107) 74256 35700 NT NT MetOH 3.57 Nitroaromatics [54]
Munitions site (Soil 108) 48365 14954 NT NT MetOH 2.29 Nitroaromatics [54]
Munitions site (Soil 109) 21.0 ND NT NT MetOH 0.21 Background [54]
Amended soil (sewage sludge) 153.0 NT 103.0 NT EDC NA [476]
Wood preserving (sludge/soil) ND 250410 ND 925110 DCM 528.0 Creosote, PCP [146]
Wood preserving (sludge/soil) ND NT ND 20 DCM 0.057 Background [146]
SWRI wastep ND 76150 ND 425970 DCM 210.0 Hydrocarbons [49]
COMBO wastep 19950 33120 ND 86920 DCM 410.0 Hydrocarbons [49]
Amended soil (creosote) ND 21144.0 466872.0 ND DCM 135.00 Creosote, PAHs [49]
Amended soil (creosote) ND 16369.0 220423.0 ND DCM 139.00 Creosote, PAHs [49]
Amended soil (refinery) 1336.0 22598.0 ND ND DCM 41.00 PAHs [49]
Amended soil (refinery) 146.0 8120.0 ND ND DCM 53.00 PAHs [49]
Amended soil (refinery) 86.0 4498.0 ND ND DCM 23.00 PAHs [49]
Amended soil (refinery) ND 5299.0 ND ND DCM 34.00 PAHs [49]
Amended soil (creosote) ND 13925.0 NT NT DCM PAHs PAHs: 2769 [121]
Amended soil (slop oil solids) 1890.0 537.0 NT NT DCM PAHs PAHs: 6646 [121]
Diesel contaminated soil 115800 ND ND ND DCM Hydrocarbons, PAHs PAHs: 115 [157]
Hazardous waste landfill NT 2330.0 NT 2740.0 DCM �149.0 Solvents, Metals [423]
Hazardous waste landfill NT 4110.0 NT 3650.0 DCM �149.0 Solvents, Metals [423]
Hazardous waste landfill NT 3040.0 NT 1693.0 Ac/Hex �149.0 Solvents, Metals [423]
Hazardous waste landfill NT 2960.0 NT 2200.0 Ac/Hex �149.0 Solvents, Metals [423]
Hazardous waste landfill NT 2540.0 NT 1680.0 Ac/Hex �149.0 Solvents, Metals [423]
Hazardous waste landfill NT 2533.0 NT 2180.0 Ac/DCM �149.0 Solvents, Metals [423]
Hazardous waste landfill NT 1903.0 NT 1893.0 Hex/Prop �149.0 Solvents, Metals [423]
Hazardous waste landfill NT 1645.0 NT 1313.0 DCM �149.0 Solvents, Metals [423]
Munitions site 81100 99300 6200 8750 EtOH/DMSO Nitroaromatics TNTq:
156 DNTs: 6.6
[65]r
Petrochemical plant NT 459.0 NT NT Ac/Hex 0.48 PAHss [56]
Tokyo soil (industrial) 125.0 283.0 NT NT Benz/EtOH NA PAHs BaP: 0.21 [476]
Katsushika-ku, Tokyo 278.0 959.0 NT NT Benz/EtOH NA PAHs BaP: 0.21 [476]
Munitions site 284000 56900 259000 163000 ACN NA Nitroaromatics TNT: 12200 [158]
Petrochem. Sludge amended 1 211.8 218.1 NT NT ACN NA PAHs PAHs: 473 [59]
Petrochem. Sludge amended 2 394.6 343.3 NT NT ACN NA PAHs PAHs: 946 [59]
Petrochem. Sludge amended 3 801.4 1511.2 NT NT ACN NA PAHs PAHs: 1892 [59]
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4
Appendix A. (Continued )
Site/sample description Salmomella mutagenic potency
in revertents per gram dry soil
Extraction
solventaEOM
(mg/g
dry soil)b
Suspected contaminants References
TA98
no S9
TA98
with S9
TA100
no S9
TA100
with S9
Name ppm dry weightc
Petrochem. Sludge amended 4 1922.5 4263.7 NT NT ACN NA PAHs PAHs: 3782 [59]
Aligarh Soil 2 276.0 279.0 306.0 296.0 MetOH NA [60]
Aligarh Soil 2 186.0 178.0 240.0 164.0 ACN NA [60]
Aligarh Soil 2 67.0 61.0 ND ND Acetone NA [60]
Carbon electrode1 30.3 17.5 62.1 61.4 Ac/cyclohex NA PAHs, Metals PAHs: 61.2 [63]
Carbon electrode2 74.4 25.7 137.3 124 Ac/cyclohex NA PAHs, Metals PAHs: 69.6 [63]
Carbon electrode3 7.6 4.7 16.9 ND Ac/cyclohex NA PAHs, Metals PAHs: 4.4 [63]
Carbon electrode4 57.9 22.3 148.5 ND Ac/cyclohex NA PAHs, Metals PAHs: 142.6 [63]
Carbon electrode5 9.3 5.4 14.4 15.9 Ac/cyclohex NA PAHs, Metals PAHs: 2.9 [63]
Carbon electrode6 14.2 6.7 ND 16.2 Ac/cyclohex NA PAHs, Metals PAHs: 4.1 [63]
Carbon electrode7 10.3 6.9 ND ND Ac/cyclohex NA PAHs, Metals PAHs: 9.6 [63]
Carbon electrode8 11.4 17 22.7 ND Ac/cyclohex NA PAHs, Metals PAHs: <1.3 [63]
Carbon electrode9 7.6 5.1 7.5 ND Ac/cyclohex NA PAHs, Metals PAHs: <1.0 [63]
Carbon electrode10 5.7 7.6 7.5 ND Ac/cyclohex NA PAHs, Metals PAHs: <1.0 [63]
a Ac: acetone, Hex: hexane, DCM: dichloromethane, CAN: acetonitrile, MetOH: methanol, Cyclohex: cyclohexane, EDC: ethylene dichloride, Prop: 2-
propanol, EtOH: ethanol, DMSO: dimethyl sulfoxide, DEK: diethylketone, Tol: toluene.b Extractable organic matter. Used to convert mutagenic potency values in revertents per g dry soil to revertents/mg extractable organic matter.c Contaminant concentration in mg/kg dry soil.d Data not available or applicable.e Polycyclic aromatic hydrocarbons.f Variety of pesticides applied, including 2,4,5,-T (2,4,5-trichlorophenoxyacetic acid), MCPA (4-chloro-2-methylphenoxyacetic acid) atrazine, bromophos, and
dimefox.g Data reported in revertents per g moist soil. Dry weight conversion assumed 27% water by weight (average value for soil texture).h Not detected (i.e., no significant positive response reported).i Not tested.j Nitroarenes such as 1,3- 1,6- and 1,8-DNP (dinitropyrene).k Benzo(a)pyrene.l Extracts of control soils (commercial soil, natural top soil, enriched garden soil) failed to elicit a positive response.m Polychlorinated dibenzofurans and polychlorinated dibenzodioxins.n 2,3,6-TCBA: 2,3,6-trichlorobenzeneacetic acid; ABN-X: acid/base/neutral extractables; 2-M-5-NBA: 2-methyl-5-nitrobenzamine.o Extracts of control soils (001, 101, 201, 301, 401) failed to elicit a positive response.p SWRI: storm water runoff impoundment sludge, COMBO: combined API separator waste treatment sludge.q Trinitrotoluene, dinitrotoluenes (various isomers).r Wet weight to dry weight conversion assumed 6–10% water by weight, the wilting point for a typical sandy loam.s Wet weight to dry weight conversion for PAH data assumed 12.5% water by weight, the permanent wilting point for a typical clay loam.
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Appendix B
Plant clastogenicity and mutagenicity data collected from the literature
Site/sample descriptiona Maximum
response observedbExposure mediumc Suspected contaminants References
Name Conc ntration
(ppm dry weight)d
1. Sister chromatid exchanges in Vicia faba
1.1. Industrial vicinity
Czech soil 11VM 30.1 Aqueous extract PAHs PAH 1.1 [68]
Czech soil 14VM 28.7 Aqueous extract PAHs PAH 0.8 [68]
Czech soil 4L 28.3 Aqueous extract PAHs [68]
Czech soil 4VM 28.3 Aqueous extract PAHs PAH 11.2 [68]
Czech soil 20VM 25.1 Aqueous extract PAHs PAH 0.2 [68]
Czech soil 3L 21.2 Aqueous extract PAHs [68]
Czech soil 1L 19.2 Aqueous extract PAHs [68]
Czech soil 2L 19.0 Aqueous extract PAHs [68]
2. Micronuclei in Vicia faba
2.1. Industrial vicinity
Czech soil 4L 2.7 Aqueous extract PAHs [68]
Czech soil 1L 1.7 Aqueous extract PAHs [68]
Czech soil 4VM 1.7 Aqueous extract PAHs PAH 11.2 [68]
Czech soil 3L 1.3 Aqueous extract PAHs [68]
Czech soil 2L 1.1 Aqueous extract PAHs [68]
Czech soil 14VM 1.1 Aqueous extract PAHs PAH 0.8 [68]
Czech soil 11VM 0.8 Aqueous extract PAHs PAH 1.1 [68]
Czech soil 20VM 0.6 Aqueous extract PAHs PAH 0.2 [68]
Cr contaminated soil 27.2 Aqueous extract Chromium Cr: 2 [82]
Cr contaminated soil 26.0 Aqueous extract Chromium Cr: 2 [82]
Cr contaminated soil 13.2 Aqueous extract Chromium Cr: 1 [82]
Cr contaminated soil 12.6 Aqueous extract Chromium Cr: 1 [82]
Cr contaminated soil 11.8 Aqueous extract Chromium Cr: 2 [82]
Cr contaminated soil 9.2 Aqueous extract Chromium Cr: 1 [82]
Cr contaminated soil 7.9 Aqueous extract Chromium Cr: 1 [82]
Slovakian soil 0.6 Direct contact Metals Cr: 2 .9, Pb: 69.5, As: 2940 [76]
e
s:
s:
s:
s:
s:
s:
s:
s:
.9
.8
.8
.9
.0
.1
.2
2
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6Appendix B. (Continued )
Site/sample descriptiona Maximum
response observedbExposure mediumc Suspected contaminants References
Name Concentration
(ppm dry weight)d
Arnoldstein soil 0.2 Direct contact Metals Cr: 25.1, Pb: 10779, As: 110 [76]
Brixlegg soil 0.2 Direct contact Metals Cr: 25.3, Pb: 2057, As: 0.2 [76]
Industrial soil A 7.7 Aqueous extract PAHs/Metals PAHs: 292.6, Cr: 2.2 (in leachate) [83]
Industrial soil B 3.9 Aqueous extract PAHs/Metals PAHs: 6978, Cr: 0.2 (in leachate) [83]
Flyash amended soil 4.6 Direct contact Metals Cr: 130.7, Ni: 31.4, Pb: 69.1, Zn: 78.3 [79]
Composted flyash soil 3.9 Direct contact Metals Cr: 117.8, Ni: 28.7, Pb: 56.9, Zn: 69.1 [79]
2.2. Others
Potting soil 0.2 Direct contact [76]
Saualpe soil 1.1 Direct contact Metals Cr: 56.2, Pb: 55.4, As: 1540 (geogenic) [76]
Control soil/dung mix 0.4 Direct contact Metals Cr: 12.2, Ni: 18.1, Pb: 15, Zn: 24.3 [79]
Compost soil/dung mix 0.3 Direct contact Metals Cr: 7.4, Ni: 9.4, Pb: 10.3, Zn: 20.3 [79]
3. Micronuclei in Tradescantia (clone 4430 unless otherwise noted)
3.1 Industrial Vicinity
Solid waste landfill 1 7.0 Aqueous extract Metals, organics [283]
Solid waste landfill 2 5.9 Aqueous extract Metals, organics [283]
Solid waste landfill 5 5.5 Aqueous extract Metals, organics [283]
Solid waste landfill 3 5.3 Aqueous extract Metals, organics [283]
Solid waste landfill 4 3.9 Aqueous extract Metals, organics [283]
Solid waste landfill 6 3.4 Aqueous extract Metals, organics [283]
Solid waste compost 2.8 DMSO extract Metals, organics [66]
Solid waste compost 2.7 DMSO extract Metals, organics [66]
Solid waste compost 2.1 DMSO extract Metals, organics [66]
Solid waste compost 1.1 DMSO extract Metals, organics [66]
Industrial soil A 11.1 Aqueous extract PAHs/Metals PAHs: 292.6, Cr: 2.2 (in leachate) [83]
Industrial soil B 7.5 Aqueous extract PAHs/Metals PAHs: 6978, Cr: 0.2 (in leachate) [83]
Mine tailings 6.9 Aqueous extract Metals, PAHs Cr: �114, Pb: �8200, As: �268 [81]
Mine tailings 6.4 Aqueous extract Metals, PAHs Cr: �114, Pb: �8200, As: �268 [81]
Mine tailings 6.1 Aqueous extract Metals, PAHs Cr: �114, Pb: �8200, As: �268 [81]
Mine tailings 4.9 Aqueous extract Metals, PAHs Cr: �114, Pb: �8200, As: �268 [81]
Letna soil 11.6 DMSO extract [69]
Letna soil 9.2 DMSO extract [69]
Florenc soil 8.7 DMSO extract [69]
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Florenc soil 6.9 DMSO extract [69]
Brixlegg soil 100.0 Direct contact [76]
Arnoldstein soil 72.0 Direct contact [76]
Slovakian soil 28.0 Direct contact [76]
Brixlegg soil 4.3 Aq. leachate (pH 4) [76]
Arnoldstein soil 4.4 Aq. leachate (pH 4) [76]
Slovakian soil 4.3 Aq. leachate (pH 4) [76]
Brixlegg soil 3.0 Aq. leachate (pH 7) [76]
Arnoldstein soil 3.7 Aq. leachate (pH 7) [76]
Slovakian soil 3.7 Aq. leachate (pH 7) [76]
Mitterghutten 1 8.1 Direct contact Metals Cr: 37, Pb: 9, As: 52.9, Cd: 0.3 [77]
Mitterghutten 2 12.9 Direct contact Metals Cr: 10, Pb: 48, As: 903, Cd: 0.7 [77]
Ramingstein 1 15.6 Direct contact Metals Cr: 31, Pb: 04, As: 24.5, Cd: 0.8 [77]
Ramingstein 2 5.7 Direct contact Metals Cr: 29, Pb: 3604, As: 232, Cd: 51.1 [77]
Bleiberg 1 6.4 Direct contact Metals Cr: 53, Pb: 3, As: 2.1, Cd: 0.3 [77]
Bleiberg 2 7.5 Direct contact Metals Cr: 13, Pb: 731, As: 4.4, Cd: 34.4 [77]
Bleiberg 3 9.7 Direct contact Metals Cr: 22, Pb: 199, As: 7.0, Cd: 62.1 [77]
Arnoldstein 1 4.8 Direct contact Metals Cr: 37, Pb: 8, As: 8.4, Cd: 0.6 [77]
Arnoldstein 2 9.2 Direct contact Metals Cr: 32, Pb: 013, As: 150, Cd: 31.2 [77]
Arnoldstein 3 8.2 Direct contact Metals Cr: 31, Pb: 8144, As: 106, Cd: 89.9 [77]
Meza 1 9.2 Direct contact Metals Cr: 56, Pb: 2, As: 15.0, Cd: 0.3 [77]
Meza 2 4.5 Direct contact Metals Cr: 124, Pb 694, As: 13.9, Cd: 7.4 [77]
Bitterfield 1 4.4 Direct contact Metals Cr: 27, Pb: 9, As: 16.0 [77]
Bitterfield 2 6.1 Direct contact Metals Cr: 214, Pb 295, As: 62 [77]
Carbon electrode 1 6.2 Aqueous extract Metals/PAHs Pb: 483.7, Z : 655.6, PAHs: 61.2 [63]
Carbon electrode 2 1.9 Aqueous extract Metals/PAHs Pb: 130.7, Z : 432.8, PAHs: 69.6 [63]
Carbon electrode 3 15.3 Aqueous extract Metals/PAHs Pb: 62.2, Zn 100.6, PAHs: 4.4 [63]
Carbon electrode 4 9.2 Aqueous extract Metals/PAHs Pb: 136.5, Z : 609.1, PAHs: 142.6 [63]
Carbon electrode 5 5.9 Aqueous extract Metals/PAHs Pb: 35.0, Zn 80.0, PAHs: 2.9 [63]
Carbon electrode 6 3.8 Aqueous extract Metals/PAHs Pb: 73.3, Zn 162.5, PAHs: 4.1 [63]
Carbon electrode 7 3.7 Aqueous extract Metals/PAHs Pb: 145.6, Z : 374.0, PAHs: 9.6 [63]
Carbon electrode 8 9.5 Aqueous extract Metals/PAHs Pb: 49.4, Zn 88.9, PAHs: <1.3 [63]
Carbon electrode 9 5.9 Aqueous extract Metals/PAHs Pb: 68.1, Zn 107.0, PAHs: <1.0 [63]
Carbon electrode 10 10.3 Aqueous extract Metals/PAHs Pb: 48.2, Zn 72.3, PAHs: <1.0 [63]
5
1
1
1
8
1
4
7
4
2
5
:
3
:
n
n
:
n
:
:
n
:
:
:
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8Appendix B. (Continued )
Site/sample descriptiona Maximum
response observedbExposure mediumc Suspected contaminants References
Name Concentration
(ppm dry weight)d
3.2. Heavily contaminated sites (e.g., hazardous waste landfill, superfund sites, etc.)
Creosote contamination 43.0 Aqueous extract Creosote, PAHs PAHs: 5749 [120]
Waste compost 8.4 DMSO extract Metals, organics [66]
Waste compost 6.3 EtOH extract Metals, organics [66]
Waste compost 6.2 DMSO extract Metals, organics [66]
Waste compost 5.8 DMSO extract Metals, organics [66]
Waste compost 5.3 DMSO extract Metals, organics [66]
Waste compost 5.3 DMSO extract Metals, organics [66]
Waste compost 5.2 Aqueous extract Metals, organics [66]
Waste compost 5.1 EtOH extract Metals, organics [66]
Waste compost 4.2 DMSO extract Metals, organics [66]
Superfund Plot 3B 23.8 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot NC1 19.3 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 2 17.9 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 3A 14.3 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 4A 11.4 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 3 10.8 Direct contact Pesticides DDT: up to 1200 [78]
Superfund Plot 4 9.8 Direct contact Pesticides DDT: up to 1200 [78]
Superfund Plot 5 9.6 Direct contact Pesticides DDT: up to 1200 [78]
Superfund Plot 1 9.3 Direct contact Pesticides DDT: up to 1200 [78]
Superfund Plot 1B 8.3 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 2A 7.9 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 2B 6.7 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 5A 5.7 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 5B 5.1 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 1A 4.7 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Superfund Plot 4B 3.5 Direct contact (slurry) Pesticides DDT: up to 1200 [78]
Rubber factory mire 7.3 Aqueous extract Organics [286]
Waste site A (Pavia) 2.5 Aqueous extract Organics [178]
Waste site B (Pavia) 5.5 Aqueous extract Organics [178]
Waste site D (Pavia) 5.9 Aqueous extract Organics [178]
3.3. Sites contaminated with mutagenic metals of geogenic origin
Saualpe soil 77.0 Direct contact Metals Cr: 56.2, Pb: 55.4, As: 1540 [76]
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Saualpe soil 5.6 Aq. leachate (pH 4) Metals Cr: 56.2, Pb: 55.4, As: 1540 [76]
Saualpe soil 3.1 Aq. leachate (pH 7) Metals Cr: 56.2, Pb: 55.4, As: 1540 [76]
Feistritz 1 11.0 Direct contact Metals Cr: 26, Pb: 34, As: 36, Cd: 0.1 [77]
Feistritz 2 16.3 Direct contact Metals Cr: 67, Pb: 54, As: 150, Cd: 0.2 [77]
3.4. Agricultural sites
Agricultural soil 6 9.1 DMSO extract Metals, organics [67]
Agricultural soil 5 5.4 DMSO extract Metals, organics [67]
Agricultural soil 4 5.4 DMSO extract Metals, organics [67]
Agricultural soil 3 5.8 DMSO extract Metals, organics [67]
Agricultural soil 2 5.2 DMSO extract Metals, organics [67]
Agricultural soil 1 3.6 DMSO extract Metals, organics [67]
MPC soil 15.2 DMSO extract Pesticides [252]
MPC soil 8.2 Aqueous extract Pesticides [252]
Vienna 1 9.6 Aqueous extract Pesticides [252]
Vienna 2 8.8 Aqueous extract Pesticides [252]
Vienna 3 7.8 Aqueous extract Pesticides [252]
Monroe soil 11.9 DMSO extract Pesticides [474]
Monroe soil 6.0 Aqueous extract Pesticides [474]
Allison soil 2.7 DMSO extract Pesticides [474]
Allison soil 3.4 Aqueous extract Pesticides [474]
Sprayed soil (high) 8.4 Aqueous extract Captan, diazinon, etc. [85]
Sprayed soil (high) 5.4 Aqueous extract Captan, diazinon, etc. [85]
Sprayed soil (medium) 7.9 Aqueous extract Captan, diazinon, etc. [85]
Sprayed soil (medium) 8.4 Aqueous extract Captan, diazinon, etc. [85]
Sprayed soil (low) 5.7 Aqueous extract Captan, diazinon, etc. [85]
Sprayed soil (low) 4.4 Aqueous extract Captan, diazinon, etc. [85]
Untertiefenbach 6.0 Direct contact Metals As: 1, Pb: 29, Cr: 43, Cd: 0.3 [77]
Reisenberg 6.3 Direct contact Metals As: 4, Pb: 37, Cr: 75, Cd: 0.7 [77]
3.5. Reference (control) sites
Karlovka soil 6.6 DMSO extract [69]
Karlovka soil 4.4 DMSO extract [69]
Control soil 8.7 Direct contact [78]
Control soil 7.7 Direct contact [78]
Control soil 7.3 Direct contact (slurry) [78]
Control soil 7.1 Direct contact [78]
Control soil 4.4 Direct contact [78]
Potting soil 6.0 Direct contact [76]
4.
4.
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0Appendix B. (Continued )
Site/sample descriptiona Maximum
response observedbExposure mediumc Suspected contaminants References
Name Concentration
(ppm dry weight)d
Potting soil 4.3 Aq. leachate (pH 4) [76]
Potting soil 4.1 Aq. leachate (pH 7) [76]
APF soil 2.7 Aqueous extract [474]
Control soil 4.0 Aqueous extract [85]
Control soil 4.9 Aqueous extract [286]
Potting soil 1 4.5 Direct contact [77]
Potting soil 2 6.4 Direct contact [77]
Potting soil 3 2.8 Direct contact [77]
Waste control (Pavia) 1.5 Aqueous extract [178]
4. Micronuclei in Allium cepa
4.1. Industrial vicinity
Industrial soil A 6.2 Aqueous extract PAHs/Metals PAHs: 292.6, Cr: 2.2 (in leachate) [83]
Industrial soil B 2.7 Aqueous extract PAHs/Metals PAHs: 6978, Cr: 0.2 (in leachate) [83]
5. Anaphase aberrations in Alliu cepa root tips
5.1. Industrial vicinity
Solid waste landfill 6.3 Aqueous extract Metals, organics [283]
Solid waste landfill 4.0 Aqueous extract Metals, organics [283]
Solid waste landfill 3.3 Aqueous extract Metals, organics [283]
Solid waste landfill 3.0 Aqueous extract Metals, organics [283]
Solid waste landfill 1.3 Aqueous extract Metals, organics [283]
Waste compost 2.0 DMSO extract Metals, organics [66]
Waste compost 0.7 DMSO extract Metals, organics [66]
Chernobyl soil 1 3.4 Direct contact 137Cs, 232Th, 90Sr 137Cs: 145 Bq/kg [257]
Chernobyl soil 2 3.1 Direct contact 137Cs, 232Th, 90Sr 137Cs: 188 Bq/kg [257]
Chernobyl soil 3 6.7 Direct contact 137Cs, 232Th, 90Sr 137Cs: 575 Bq/kg [257]
Chernobyl soil 4 6.0 Direct contact 137Cs, 232Th, 90Sr 137Cs: 582 Bq/kg [257]
Bhopal soil 1 5.8 Aqueous extract MICe [477]
Bhopal soil 2 1.6 Aqueous extract MIC [477]
5.2. Heavily contaminated sites (e.g., hazardous waste landfill, Superfund sites, etc.)
Waste compost 2.2 DMSO extract Metals, organics [66]
Waste compost 1.6 DMSO extract Metals, organics [66]
Chernobyl soil 5 13.7 Direct contact 137Cs, 232Th, 90Sr 137Cs: 2287 Bq/kg [257]
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Chernobyl soil 6 14.9 Direct contact 137Cs, 232Th, 90Sr 137Cs: 2543 Bq/kg [257]
Chernobyl soil 7 23.8 Direct contact 137Cs, 232Th, 90Sr 137Cs: 6549 Bq/kg [257]
PCB contaminated soil 3.2 Aqueous extract PCBs PCBs: 144 [254]
5.3. Agricultural sites
Agricultural soil 2 0.6 DMSO extract Metals, organics [67]
Agricultural soil 3 1.8 DMSO extract Metals, organics [67]
Agricultural soil 4 0.8 DMSO extract Metals, organics [67]
Agricultural soil 5 2.2 DMSO extract Metals, organics [67]
Agricultural soil 6 3.0 DMSO extract Metals, organics [67]
Vienna 1 1.6 Aqueous extract Pesticides [78]
Vienna 2 1.7 Aqueous extract Pesticides [78]
Vienna 3 1.4 Aqueous extract Pesticides [78]
MPC soil 2.1 Aqueous extract Pesticides [78]
Ukrainian soil 37.1 6.6 Direct contact [74]
Ukrainian soil 37 6.0 Direct contact [74]
Ukrainian soil 18 5.2 Direct contact [74]
Ukrainian soil 30 4.3 Direct contact [74]
Ukrainian soil 16 3.6 Direct contact [74]
Ukrainian soil 14 3.6 Direct contact [74]
Ukrainian soil 2 3.4 Direct contact [74]
Ukrainian soil 27 3.3 Direct contact [74]
Ukrainian soil 34 3.1 Direct contact [74]
Ukrainian soil 4 3.0 Direct contact [74]
Ukrainian soil 15 2.7 Direct contact [74]
Ukrainian soil 7 2.7 Direct contact [74]
Ukrainian soil 11 2.5 Direct contact [74]
Ukrainian soil 33 2.3 Direct contact [74]
Ukrainian soil 37.2 2.3 Direct contact [74]
Ukrainian soil 12 2.3 Direct contact [74]
Ukrainian soil 32 2.2 Direct contact [74]
Ukrainian soil 35 2.1 Direct contact [74]
Ukrainian soil 10 1.9 Direct contact [74]
Ukrainian soil 21 1.9 Direct contact [74]
Ukrainian soil 20 1.9 Direct contact [74]
Ukrainian soil 39 1.8 Direct contact [74]
Ukrainian soil 8 1.6 Direct contact [74]
Ukrainian soil 37.3 1.4 Direct contact [74]
Ukrainian soil 5 1.3 Direct contact [74]
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2Appendix B. (Continued )
Site/sample descriptiona Maximum
response observedbExposure mediumc Suspected contaminants References
Name Concentration
(ppm dry weight)d
Ukrainian soil 36 1.1 Direct contact [74]
Ukrainian soil 3 1.0 Direct contact [74]
Ukrainian soil 40 1.0 Direct contact [74]
Ukrainian soil 19 1.0 Direct contact [74]
Ukrainian soil 31 0.8 Direct contact [74]
Ukrainian soil 1 0.6 Direct contact [74]
Uzbekistan Soil 1 3.8 Direct contact [75]
Uzbekistan Soil 2 2.9 Direct contact [75]
Uzbekistan Soil 3 4.9 Direct contact [75]
Uzbekistan Soil 4 0.8 Direct contact [75]
Uzbekistan Soil 5 5.1 Direct contact [75]
Uzbekistan Soil 6 6.1 Direct contact [75]
Uzbekistan Soil 7 1.7 Direct contact [75]
Uzbekistan Soil 8 2.6 Direct contact [75]
Uzbekistan Soil 9 3.8 Direct contact [75]
Uzbekistan Soil 10 1.0 Direct contact [75]
Uzbekistan Silt 1 5.3 Direct contact [75]
Uzbekistan Silt 2 4.2 Direct contact [75]
Uzbekistan Silt 3 2.5 Direct contact [75]
Uzbekistan Silt 4 2.7 Direct contact [75]
Uzbekistan Silt 5 5.4 Direct contact [75]
Uzbekistan Silt 6 6.9 Direct contact [75]
Uzbekistan Silt 7 4.2 Direct contact [75]
Uzbekistan Silt 8 4.0 Direct contact [75]
Uzbekistan Silt 9 4.2 Direct contact [75]
Uzbekistan Silt 10 3.0 Direct contact [75]
Uzbekistan Soil 11 4.6 Direct contact [75]
Uzbekistan Soil 12 5.3 Direct contact [75]
Uzbekistan Soil 13 2.8 Direct contact [75]
Uzbekistan Soil 14 2.6 Direct contact [75]
Uzbekistan Soil 15 5.0 Direct contact [75]
Uzbekistan Soil 16 4.0 Direct contact [75]
Uzbekistan Soil 17 4.1 Direct contact [75]
Uzbekistan Soil 18 6.4 Direct contact [75]
Uzbekistan Soil 19 3.3 Direct contact [75]
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Uzbekistan Soil 20 4.2 Direct contact [75]
Uzbekistan Silt 11 3.6 Direct contact [75]
Uzbekistan Silt 12 4.4 Direct contact [75]
Uzbekistan Silt 13 2.1 Direct contact [75]
Uzbekistan Silt 14 4.0 Direct contact [75]
Uzbekistan Silt 15 5.6 Direct contact [75]
Uzbekistan Silt 16 4.5 Direct contact [75]
Uzbekistan Silt 17 6.2 Direct contact [75]
Uzbekistan Silt 18 4.2 Direct contact [75]
Uzbekistan Silt 19 6.5 Direct contact [75]
Uzbekistan Silt 20 4.2 Direct contact [75]
5.4. Reference (control) sites
Uzbekistan control 1 1.6 Direct contact [75]
Uzbekistan control 2 1.8 Direct contact [75]
Chernobyl control soil 1.6 Direct contact [257]
Pesticide-free control 0.8 Aqueous extract [252]
PCB Reference soil 0.6 Aqueous extract [254]
6. Gametic mutations in Arabidopsis thaliana (Muller embryo test)
6.1. Industrial vicinity
Czech soil 20VM 35.0 DCM extract PAHs PAHs: 0.2 [68]
Czech soil 4VM 34.5 DCM extract PAHs PAHs: 11.2 [68]
Czech soil 4VM 34.4 Aqueous extract PAHs PAHs: 11.2 [68]
Czech soil 11VM 28.6 Aqueous extract PAHs PAHs: 1.1 [68]
Czech soil 20VM 27.1 Aqueous extract PAHs [68]
Czech soil 11VM 27.1 DCM extract PAHs PAHs: 1.1 [68]
Czech soil 2VM 24.6 DCM extract PAHs [68]
Czech soil 4L 24.3 Aqueous extract PAHs [68]
Czech soil 14VM 20.3 DCM extract PAHs PAHs: 0.8 [68]
Czech soil 14VM 19.8 Aqueous extract PAHs PAHs: 0.8 [68]
Czech soil 3L 9.4 DCM extract PAHs [68]
Czech soil 2L 6.4 DCM extract PAHs [68]
Czech soil 1L 5.9 DCM extract PAHs [68]
Czech soil 4L 5.2 DCM extract PAHs [68]
Czech soil 3L 3.5 Aqueous extract PAHs [68]
Czech soil 1L 1.6 Aqueous extract PAHs [68]
Czech soil 2L 0.7 Aqueous extract PAHs [68]
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4Appendix B. (Continued )
Site/sample descriptiona Maximum
response observedbExposure mediumc Suspected contaminants References
Name Concentration
(ppm dry weight)d
6.2. Heavily contaminated sites (e.g., hazardous waste landfill, Superfund sites, etc.)
Chernobyl soil 1 8.2 In situ Radioactive Cs 134Cs + 137Cs: 217 Bq/kg [258]
Chernobyl soil 1 20.2 In situ Radioactive Cs 134Cs + 137Cs: 1025 Bq/kg [258]
Chernobyl soil 1 51.6 In situ Radioactive Cs 134Cs + 137Cs: 2529 Bq/kg [258]
7. Zea mays — pollen mutations at waxy locusf
7.1. Industrial vicinity
Wood river #1 (1978) 72.5 Direct contact Petrochemicals [71]
Wood river #1 (1979) 41.6 Direct contact Petrochemicals [71]
Beaumont TX #1 29.9 Direct contact Petrochemicals [71]
Beaumont TX #2 29.0 Direct contact Petrochemicals [71]
Wood river #1 (1979) 5.0 Direct contact Petrochemicals [71]
Wood river #1 (1980) 20.0 Direct contact Petrochemicals [71]
Wood river #2 (1978) 22.8 Direct contact Petrochemicals [71]
Smelter 1978 (0.3 km) 34.6 Direct contact Lead [70]
Smelter 1976 (1.7 km) 28.3 Direct contact Lead [70]
Smelter 1976 (0.3 km) 22.0 Direct contact Lead [70]
Smelter 1978 (1.7 km) 12.1 Direct contact Lead [70]
Smelter 1977 (3.2 km) 8.6 Direct contact Lead [70]
Smelter 1977 (1.7 km) 7.3 Direct contact Lead [70]
Smelter 1977 (0.3 km) 6.7 Direct contact Lead [70]
Smelter 1978 (3.2 km) 3.7 Direct contact Lead [70]
Sludge amended soil 18.5 Direct contact Metals, organics [72]g
7.2. Agricultural sites
Moderate pesticide 6.0 Direct contact Captan [84]h
Moderate pesticide 5.9 Direct contact Captan + diazinon [84]h
Broadcast pesticide 7.7 Direct contact Above + cyanazine,
metalochlor, chlorpyrifos,
and lindane
[84]h
Broadcast pesticide 8.5 Direct contact Above + cyanazine,
metalochlor, chlorpyrifos,
and lindane
[84]h
Herbicide application 4.21 Direct contact Alachlor: 6.00 kg/ha [270]
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Herbicide application 3.82 Direct contact Alachlor + dicamba: 2.34 + 0.56 kg/ha [270]
Herbicide application 8.53 Direct contact Atrazine: 3.84 kg/ha [270]
Herbicide application 9.05 Direct contact Bifenox: 2.24 kg/ha [270]
Herbicide application 5.65 Direct contact Bifenox+alachlor: 1.40 + 2.29 kg/ha (means) [270]
Herbicide application 5.72 Direct contact Butylate: 7.20 kg/ha [270]
Herbicide application 8.25 Direct contact Butylate + atrazine: 4.80 + 1.92 kg/ha [270]
Herbicide application 5.90 Direct contact Butylate + cyanazine: 4.80 + 2.24 kg/ha [270]
Herbicide application 21.51 Direct contact Cyanazine: 4.19 kg/ha (mean) [270]
Herbicide application 2.43 Direct contact Dicamba: 0.56 kg/ha [270]
Herbicide application 4.31 Direct contact Eradicane: 7.20 kg/ha [270]
Herbicide application 14.20 Direct contact Eradicane + atrazine: 40 kg/ha [270]
Herbicide application 8.61 Direct contact Eradicane + cyanazine: 3.60 + 2.40 kg/ha [270]
Herbicide application 2.84 Direct contact Metolachlor: 8.40 kg/ha [270]
Herbicide application 12.27 Direct contact Metolachlor + atrazine: 3.00 + 2.40 kg/ha [270]
Herbicide application 10.84 Direct contact Metolachlor + cyanazine: 4.80 + 4.80 kg/ha [270]
Herbicide application 7.77 Direct contact Metalachlor + dicamba: 3.00 + 0.60 kg/ha [270]
Herbicide application 4.64 Direct contact Procyazine: 3.58 kg/ha [270]
Herbicide application 9.83 Direct contact Procyazine + metolachlor: 2.24 + 2.24 kg/ha [270]
Herbicide application 3.20 Direct contact Propachlor: 3.36 kg/ha [270]
Herbicide application 6.29 Direct contact Propachlor + cyanazine: 4.80 + 2.24 kg/ha [270]
Herbicide application 27.81 Direct contact SD50093: 4.64 kg/ha (mean) [270]
Herbicide application 10.86 Direct contact Simazine: 3.80 kg/ha [270]
Insecticide application 6.15 Direct contact Carbofuran: 2.24 kg/ha [271]
Insecticide application 9.72 Direct contact Chlordane: 2.24 kg/ha [271]
Insecticide application 2.74 Direct contact Chlorpyrifos: 2.24 kg/ha [271]
Insecticide application 6.73 Direct contact Curacron: 2.24 kg/ha [271]
Insecticide application 7.92 Direct contact Ethoprop: 2.24 kg/ha [271]
Insecticide application 6.14 Direct contact Fonofos: 2.24 kg/ha [271]
Insecticide application 10.40 Direct contact Heptachlor: 1.12 kg/ha [271]
Insecticide application 6.85 Direct contact Metham: 2.24 kg/ha [271]
Insecticide application 4.08 Direct contact Phorate: 2.24 kg/ha [271]
Insecticide application 7.30 Direct contact Terbufos: 2.24 kg/ha [271]
7.3. Reference (control) sites
Reference soil 5.5 Direct contact [71]
Reference soil 2.6 Direct contact [71]
Reference soil 2.6 Direct contact [71]
Smelter reference 1976 12.9 Direct contact [70]
Reference soil 8.3 Direct contact [70]
Reference soil 6.5 Direct contact [70]
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6Appendix B. (Continued )
Site/sample descriptiona Maximum
response observedbExposure mediumc Suspected contaminants References
Name Concentration
(ppm dry weight)d
Smelter reference 1977 4.6 Direct contact [70]
Reference 3.1 Direct contact [84]
No pesticide 4.4 Direct contact [84]
Reference soil 1.8 Direct contact [72]g
Reference soil 4.1 Direct contact [270]
Reference soil 4.0 Direct contact [271]
8. Stamen hair mutations inTradescantia sp.i
8.1. Industrial vicinity
Beaumont TX 1.2 7.2 Direct contact Petrochemicals [71]
Beaumont TX 1.4 6.0 Direct contact Petrochemicals [71]
Beaumont TX 1.1 5.5 Direct contact Petrochemicals [71]
Beaumont TX 1.3 5.3 Direct contact Petrochemicals [71]
Beaumont TX 1.11 5.3 Direct contact Petrochemicals [71]
Beaumont TX 1.5 5.1 Direct contact Petrochemicals [71]
Beaumont TX 1.10 4.7 Direct contact Petrochemicals [71]
Beaumont TX 1.6 4.6 Direct contact Petrochemicals [71]
Beaumont TX 1.7 4.5 Direct contact Petrochemicals [71]
Beaumont TX 1.8 3.4 Direct contact Petrochemicals [71]
Beaumont TX 1.9 3.4 Direct contact Petrochemicals [71]
Wood river 4.8 Direct contact Petrochemicals [71]
Granite city 3.9 Direct contact Petrochemicals [71]
Smelter 1977 (0.3 km) 4.9 Direct contact Lead [70]
Smelter 1977 (1.7 km) 4.8 Direct contact Lead [70]
Smelter 1978 (1.7 km) 4.6 Direct contact Lead [70]
Smelter 1977 (3.2 km) 4.4 Direct contact Lead [70]
Smelter 1977 (0.3 km) 4.1 Direct contact Lead [70]
Smelter 1977 (3.2 km) 3.9 Direct contact Lead [70]
Smelter 1978 (7.4 km) 3.9 Direct contact Lead [70]
Smelter 1977 (1.7 km) 3.7 Direct contact Lead [70]
Smelter 1978 (3.2 km) 3.5 Direct contact Lead [70]
Smelter 1978 (7.4 km) 3.4 Direct contact Lead [70]
Smelter 1978 (0.3 km) 3.0 Direct contact Lead [70]
Smelter 1977 (1.7 km) 2.9 Direct contact Lead [70]
Smelter 1978 (3.2 km) 2.9 Direct contact Lead [70]
Smelter 1977 (0.3 km) 2.8 Direct contact Lead [70]
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Smelter 1977 (3.2 km) 2.7 Direct contact Lead [70]
Smelter 1976 (1.7 km) 2.6 Direct contact Lead [70]
Smelter 1978 (1.7 km) 2.6 Direct contact Lead [70]
Smelter 1976 (0.3 km) 2.2 Direct contact Lead [70]
Smelter 1976 (0.3 km) 2.1 Direct contact Lead [70]
Bikini Islands B1-6 2.9 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B1-8 2.6 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B1-4 2.6 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B1-9 2.6 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B1-2 2.6 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B1-7 2.5 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B1-3 2.4 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B1-5 2.4 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B1-1 1.8 Direct contact Radionuclides 137Cs: 688 Bq/kg [73]j
Bikini Islands B2-5 2.2 Direct contact Radionuclides [73]j
Bikini Islands B2-1 2.3 Direct contact Radionuclides [73]j
Bikini Islands B2-2 2.2 Direct contact Radionuclides [73]j
Bikini Islands B2-6 2.1 Direct contact Radionuclides [73]j
Bikini Islands B2-8 2.1 Direct contact Radionuclides [73]j
Bikini Islands B2-9 2.0 Direct contact Radionuclides [73]j
Bikini Islands B2-7 1.9 Direct contact Radionuclides [73]j
Bikini Islands B2-4 1.7 Direct contact Radionuclides [73]j
Bikini Islands B2-3 1.6 Direct contact Radionuclides [73]j
Letna soil 1.9 DMSO extract [120]
Letna soil 1.2 DMSO extract [120]
Florenc soil 1.4 DMSO extract [120]
Florenc soil 0.9 DMSO extract [120]
Mine tailings 5.8 Aqueous extract Metals, PAHs Cr: �114, b: �8200, As: �268 [81]
Mine tailings 4.1 Aqueous extract Metals, PAHs Cr: �114, b: �8200, As: �268 [81]
Mine tailings 4.1 Aqueous extract Metals, PAHs Cr: �114, b:�8200, As: �268 [81]
Mine tailings 3.7 Aqueous extract Metals, PAHs Cr: �114, b: �8200, As: �268 [81]
Waste compost 2.8 DMSO extract Metals, organics [66]
Waste compost 1.8 DMSO extract Metals, organics [66]
Waste compost 1.7 DMSO extract Metals, organics [66]
Waste compost 1.7 DMSO extract Metals, organics [66]
Solid waste landfill 3.5 Aqueous extract Metals, organics [283]
Solid waste landfill 3.4 Aqueous extract Metals, organics [283]
Solid waste landfill 3.3 Aqueous extract Metals, organics [283]
0
0
0
0
0
0
0
0
0
P
P
P
P
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
32
8Appendix B. (Continued )
Site/sample descriptiona Maximum
response observedbExposure mediumc Suspected contaminants References
Name Concentration
(ppm dry weight)d
Solid waste landfill 3.3 Aqueous extract Metals, organics [283]
Solid waste landfill 2.9 Aqueous extract Metals, organics [283]
8.2. Heavily contaminated sites (e.g., hazardous waste landfill, Superfund sites, etc.)
Waste compost 2.4 DMSO extract Metals, organics [66]
Waste compost 1.8 DMSO extract Metals, organics [66]
Waste compost 1.8 DMSO extract Metals, organics [66]
Waste compost 1.7 DMSO extract Metals, organics [66]
8.3. Agricultural sites
Agricultural soil 6 7.2 DMSO extract Metals, organics [67]
Agricultural soil 5 5.2 DMSO extract Metals, organics [67]
Agricultural soil 4 5.1 DMSO extract Metals, organics [67]
Agricultural soil 2 3.6 DMSO extract Metals, organics [67]
Agricultural soil 1 3.6 DMSO extract Metals, organics [67]
Agricultural soil 3 3.4 DMSO extract Metals, organics [67]
Ukrainian soil 2.4 Direct contact [74]k
Ukrainian soil 2.0 Direct contact [74]k
Ukrainian soil 1.6 Direct contact [74]k
Ukrainian soil 1.5 Direct contact [74]k
Ukrainian soil 0.9 Direct contact [74]k
Ukrainian soil 0.3 Direct contact [74]k
Uzbekistan Soil 1 3.0 Direct contact [75]k
Uzbekistan Soil 2 1.6 Direct contact [75]k
Uzbekistan Soil 3 1.5 Direct contact [75]k
Uzbekistan Soil 4 1.2 Direct contact [75]k
Uzbekistan Soil 5 1.5 Direct contact [75]k
8.4. Reference (control) sites
Karlovka soil 1.3 DMSO extract [69]
Karlovka soil 1.2 Aqueous extract [69]
Control soil 2.3 Direct contact [73]j
Control soil 1.9 Direct contact [73]j
Control soil 1.8 Direct contact [73]j
Control soil 1.8 Direct contact [73]j
Control soil 1.7 Direct contact [73]j
P.A
.W
hite,
L.D
.C
laxto
n/M
uta
tion
Resea
rch5
67
(20
04
)2
27
–3
45
32
9
Control soil 1.7 Direct contact [73]j
Control soil 1.7 Direct contact [73]j
Control soil 1.6 Direct contact [73]j
Control soil 1.6 Direct contact [73]j
Vidor TX 3.9 Direct contact [71]
Control soil 1.3 Direct contact [71]
Columbia MO 1.3 Direct contact [71]
Smelter control 1977 3.5 Direct contact [70]
Smelter control 1977 3.4 Direct contact [70]
Smelter control 1978 2.4 Direct contact [70]
Smelter control 1976 2.0 Direct contact [70]
Control soil 3.2 Direct contact [70]
Control soil 2.8 Direct contact [70]
Control soil 2.6 Direct contact [70]
Control soil 2.5 Direct contact [70]
Control soil 1.7 Direct contact [70]
Control soil 1.7 Direct contact [70]
aSites are divided into control/reference sites, sites that are primarily agricultural, sites selected for their proxim y to large industries, heavily contaminated
waste disposal and/or containment sites, and sites containing mutagenic metals of geologic origin.b Units are as follows: Tradescantia micronucleus assay — micronuclei per 100 tetrads; Vicia faba micronucleu assay — micronuclei per 1000 cells; sister
chromatid exchanges in Vicia faba — SCEs per cell; chromosomal aberrations in Allium cepa — total aberrations r 100 cells; stamen hair mutation assay in
Tradescantia — mutants per 1000 hairs; waxy mutants in Zea mays —waxy locus mutations in mutants �105; abidopsis gametic mutations — mutation
frequency in %.c Includes aqueous extracts, aqueous leachates, diluted organic extracts, direct contact soil exposures, and exp sures to aqueous soil slurries.d Concentration in the original soil sample, unless otherwise noted.e Methylisocyanate and related break-down products.f Reverse mutation at the wx-C allele in inbred line W22 unless otherwise noted.g Reverse mutation at the wx-C or wx-90 allele in inbred line M14.h Forward waxy locus mutation in Early-Early Synthetic variety.i Stamen hair mutations in Tradescantia clone 4430 unless otherwise noted.j Stamen hair mutations in Tradescantia clone BNL02 (Brookhaven National Laboratory 02).k Stamen hair mutations in an undescribed isolate of Tradescantia poludosa.
it
s
pe
Ar
o
P.A. White, L.D. Claxton / Mutation Research 567 (2004) 227–345330
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